metadata: run_metadata: 3,059 rows where experiment.library_layout = "SINGLE", experiment.library_source = "TRANSCRIPTOMIC SINGLE CELL" and experiment.platform = "ILLUMINA"

3,059 rows where experiment.library_layout = "SINGLE", experiment.library_source = "TRANSCRIPTOMIC SINGLE CELL" and experiment.platform = "ILLUMINA"

descending

Link	rowid ▼	run.accession	experiment.accession	sample.accession	study.accession	bioproject	study.title	study.alias	study.type	study.abstract	study.attributes	study.PMIDs	sample.description	sample.title	sample.alias	sample.centername	sample.attributes	GEOsample.title	GEOsample.dataprocessing	GEOsample.source	GEOsample.extractprotocol	GEOsample.characteristics	GEOsample.accession	experiment.title	experiment.alias	experiment.library_name	experiment.design_description	experiment.library_construction_protocol	experiment.attributes	experiment.library_strategy	experiment.library_source	experiment.library_selection	experiment.library_layout	experiment.platform	experiment.instrument_model	experiment.study_ref	run.title	run.attributes	run.filename	run.semantic_name	run.total_bases	run.total_spots	run.alias	run.read_lengths	run.base_counts	run.r1_length	run.r2_length	run.r3_length	run.r4_length	run.Acount	run.Ccount	run.Gcount	run.Tcount	run.Ncount	run.experiment	run.pool_member	submission.accession	submission.srasource	submission.bioprojectsource	seqdetective.n_mates	seqdetective.mapping_rate.mate1	seqdetective.nofeature_rate.mate1	seqdetective.sparsity.mate1	seqdetective.pos_strand_rate.mate1	seqdetective.readlen.mate1	seqdetective.judgement.mate1	seqdetective.judgement.reason	platform_family	instrument_generation	read_bias	selection_class	prep_kit	sc_or_bulk	tech_class	technology	submission.bioprojectsource.country	earliest_date	devstage_curation	devstage_curation_coarse	tissue_curation	tissue_curation_coarse
9898	9898	ERR4194114	ERX4155254	ERS4601292	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 2	SAMEA6873729	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873729\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 2\|age:8\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 2\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 2 s	Sample 2 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:8\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA8h_1_S2_L001.bam	bam	10087287534.0	99874134.0	E MTAB 9193:cDNA8h 1 S2 L001	0:101	A:2892833391;C:2022466595;G:2198646699;T:2962268446;N:11072403	101				2892833391	2022466595	2198646699	2962268446	11072403	ERX4155254	ERS4601292	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93609	0.12902	0.82158	0.5062	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9899	9899	ERR4194115	ERX4155254	ERS4601292	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 2	SAMEA6873729	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873729\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 2\|age:8\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 2\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 2 s	Sample 2 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:8\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA8h_1_S2_L002.bam	bam	10191911414.0	100910014.0	E MTAB 9193:cDNA8h 1 S2 L002	0:101	A:2923216190;C:2043975715;G:2221734566;T:2992798366;N:10186577	101				2923216190	2043975715	2221734566	2992798366	10186577	ERX4155254	ERS4601292	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93445	0.13038	0.824	0.49774	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9900	9900	ERR4194112	ERX4155253	ERS4601291	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 1	SAMEA6873728	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873728\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 1\|age:6\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 1\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 1 s	Sample 1 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:6\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA6h_1_S1_L001.bam	bam	7181772357.0	71106657.0	E MTAB 9193:cDNA6h 1 S1 L001	0:101	A:2074032710;C:1415439071;G:1544274166;T:2140112533;N:7913877	101				2074032710	1415439071	1544274166	2140112533	7913877	ERX4155253	ERS4601291	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.9297	0.12156	0.8117	0.51457	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9901	9901	ERR4194113	ERX4155253	ERS4601291	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 1	SAMEA6873728	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873728\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 1\|age:6\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 1\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 1 s	Sample 1 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:6\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA6h_1_S1_L002.bam	bam	7256542253.0	71846953.0	E MTAB 9193:cDNA6h 1 S1 L002	0:101	A:2096163385;C:1430325700;G:1560530043;T:2162265616;N:7257509	101				2096163385	1430325700	1560530043	2162265616	7257509	ERX4155253	ERS4601291	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.92882	0.12127	0.81162	0.51457	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9902	9902	ERR4194128	ERX4155256	ERS4601294	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 4	SAMEA6873731	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873731\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 4\|age:13\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 4\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 4 s	Sample 4 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:13\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA13h_1_control_S1_L001.bam	bam	10785974123.0	106791823.0	E MTAB 9193:cDNA13h 1 control S1 L001	0:101	A:3007305661;C:2241057062;G:2505494392;T:2986021199;N:46095809	101				3007305661	2241057062	2505494392	2986021199	46095809	ERX4155256	ERS4601294	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.67094	0.07571	0.92951	0.5272	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9903	9903	ERR4194129	ERX4155256	ERS4601294	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 4	SAMEA6873731	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873731\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 4\|age:13\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 4\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 4 s	Sample 4 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:13\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA13h_1_control_S1_L002.bam	bam	10595192294.0	104902894.0	E MTAB 9193:cDNA13h 1 control S1 L002	0:101	A:2971916419;C:2204229263;G:2390854035;T:2949877754;N:78314823	101				2971916419	2204229263	2390854035	2949877754	78314823	ERX4155256	ERS4601294	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.67255	0.08054	0.91583	0.52661	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9904	9904	ERR4194116	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_1_S3_L001.bam	bam	1622536412.0	16556494.0	E MTAB 9193:cDNA10h 1 S3 L001	0:98	A:491141175;C:317161060;G:354179755;T:459976409;N:78013	98				491141175	317161060	354179755	459976409	78013	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93238	0.13033	0.89132	0.47076	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9905	9905	ERR4194117	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_1_S3_L002.bam	bam	1484328776.0	15146212.0	E MTAB 9193:cDNA10h 1 S3 L002	0:98	A:450782055;C:290088574;G:322801188;T:420570801;N:86158	98				450782055	290088574	322801188	420570801	86158	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.92846	0.13128	0.89923	0.46879	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9906	9906	ERR4194118	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_1_S3_L003.bam	bam	1578833116.0	16110542.0	E MTAB 9193:cDNA10h 1 S3 L003	0:98	A:477600666;C:308260969;G:347303289;T:445467384;N:200808	98				477600666	308260969	347303289	445467384	200808	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93107	0.12965	0.91265	0.47375	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9907	9907	ERR4194119	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_1_S3_L004.bam	bam	1859564994.0	18975153.0	E MTAB 9193:cDNA10h 1 S3 L004	0:98	A:562936790;C:364057256;G:406607636;T:525811595;N:151717	98				562936790	364057256	406607636	525811595	151717	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93453	0.1266	0.87714	0.46957	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9908	9908	ERR4194120	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L001.bam	bam	1156341102.0	11799399.0	E MTAB 9193:cDNA10h 2 S3 L001	0:98	A:345857612;C:224453195;G:257551986;T:327969064;N:509245	98				345857612	224453195	257551986	327969064	509245	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93863	0.10744	0.81984	0.501	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9909	9909	ERR4194121	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L002.bam	bam	1105219696.0	11277752.0	E MTAB 9193:cDNA10h 2 S3 L002	0:98	A:331051973;C:214369903;G:246546338;T:312897773;N:353709	98				331051973	214369903	246546338	312897773	353709	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93941	0.10818	0.82211	0.50079	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9910	9910	ERR4194122	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L003.bam	bam	1105199606.0	11277547.0	E MTAB 9193:cDNA10h 2 S3 L003	0:98	A:331158619;C:214305131;G:246465559;T:312930408;N:339889	98				331158619	214305131	246465559	312930408	339889	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93767	0.10645	0.82329	0.50057	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9911	9911	ERR4194123	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L004.bam	bam	1108707712.0	11313344.0	E MTAB 9193:cDNA10h 2 S3 L004	0:98	A:331374433;C:215925150;G:247131942;T:313865174;N:411013	98				331374433	215925150	247131942	313865174	411013	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93827	0.10842	0.82031	0.49241	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9912	9912	ERR4194124	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L005.bam	bam	1110084612.0	11327394.0	E MTAB 9193:cDNA10h 2 S3 L005	0:98	A:332881960;C:215360330;G:247563319;T:313842331;N:436672	98				332881960	215360330	247563319	313842331	436672	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93827	0.10719	0.8238	0.50406	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9913	9913	ERR4194125	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L006.bam	bam	1113812728.0	11365436.0	E MTAB 9193:cDNA10h 2 S3 L006	0:98	A:333112288;C:216649152;G:248341020;T:315338848;N:371420	98				333112288	216649152	248341020	315338848	371420	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93843	0.10675	0.82079	0.49284	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9914	9914	ERR4194126	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L007.bam	bam	1119495748.0	11423426.0	E MTAB 9193:cDNA10h 2 S3 L007	0:98	A:335288339;C:217283719;G:249624836;T:316900388;N:398466	98				335288339	217283719	249624836	316900388	398466	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93711	0.10749	0.82266	0.49382	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9915	9915	ERR4194127	ERX4155255	ERS4601293	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 3	SAMEA6873730	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873730\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 3\|age:10\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 3\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 3 s	Sample 3 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:10\|Experimental Factor: RNA interference:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA10h_2_S3_L008.bam	bam	1175946688.0	11999456.0	E MTAB 9193:cDNA10h 2 S3 L008	0:98	A:351364169;C:228318193;G:261971072;T:333863943;N:429311	98				351364169	228318193	261971072	333863943	429311	ERX4155255	ERS4601293	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.93851	0.10755	0.82158	0.49895	98	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9916	9916	ERR4194130	ERX4155257	ERS4601295	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 5	SAMEA6873732	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873732\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 5\|age:13\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 5\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 5 s	Sample 5 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:13\|Experimental Factor: RNA interference:morpholino mediated Gata5/6 knockdown	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA13h_2_gata56KD_S2_L001.bam	bam	7791309377.0	77141677.0	E MTAB 9193:cDNA13h 2 gata56KD S2 L001	0:101	A:2198242220;C:1593746618;G:1784167452;T:2181979820;N:33173267	101				2198242220	1593746618	1784167452	2181979820	33173267	ERX4155257	ERS4601295	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.66736	0.08039	0.93026	0.50796	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
9917	9917	ERR4194131	ERX4155257	ERS4601295	ERP122151	PRJEB38705	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E-MTAB-9193	Transcriptome Analysis	TgBACgata5:EGFP transgenic line was used to isolate mesendoderm cells enriched for cardiac lineage. Wild type embryos at 6 8 10 13 hpf and morpholinos injected Gata5/6 know down KD embryos at 13 hpf were pre screened for GFP signal and dissociated into single cell suspension in separate wells. GFP+ cells at each condition were isolated through fluorescent activated cell sorting FACS. Single cell cDNA libraries were constructed through the 10x Chromium Single Cell Gene Expression platform. Final sequencing was performed on an Illumina HiSeq 2500 platform Rapid Run Model. Note that the 10h single cell library was sequenced twice Named as cDNA10h 1* and cDNA10h 2* separately.	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01		Protocols: WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Sample 5	SAMEA6873732	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada	ENA FIRST PUBLIC:2020 12 31T11:11:47Z\|ENA LAST UPDATE:2020 06 04T16:16:46Z\|External Id:SAMEA6873732\|INSDC center name:Developmental and Stem Cell Biology Sickkids Toronto Canada; Genetics and Genome Biology Sickkids Toronto Canada; Molecular Genetics University of Toronto Toronto Canada\|INSDC first public:2020 12 31T11:11:47Z\|INSDC last update:2020 06 04T16:16:46Z\|INSDC status:public\|Submitter Id:E MTAB 9193:Sample 5\|age:13\|broker name:ArrayExpress\|common name:zebrafish\|developmental stage:embryo stage\|genotype:TgBACgata5: EGFPpd25\|immunophenotype:GFP+\|individual:embryo pool 100 200\|organism part:mesendoderm\|sample name:E MTAB 9193:Sample 5\|scientific name:Danio rerio\|sex:mixed							Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	E MTAB 9193:Sample 5 s	Sample 5 s	Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	WT and Gata5/6KD TgBAC gata5: EGFP embryos were synchronized and dechorionated with pronase Sigma Cat# 11459643001 at desired stages. For embryos at gastrulation stages 6 hpf 8 hpf 10 hpf the dissociation was performed by incubating embryos in 200 μl calcium free Ringer solution 116 mM NaCl 2.6 mM KCl 5 mM HEPE pH 7.0 for 5 min followed by 500ul TrpLE GIBCO TrypLE Express Enzyme cat #: 12604 013 at room temperature. Dissociation was monitored under a dissecting scope and embryos were homogenized every 5min until single cell suspensions were generated. Embryos at the early segmentation stage 13 hpf were treated similarly but incubated with 500 ul of 0.25 Trypsin GIBCO Trypsin2.5% no phenol red cat#: 15090046/ EDTA instead of TrpLE for dissociation. post dissociation Cells were washed twice with DMEM 1% FBS. DAPI was added at a concentration of 5ug/ml to exclude dead cells right before the Fluorescence activated cell sorting FACS. FACS was performed on Sony SH800S Cell Sorter MoFlo XDP or MoFlo Astrios with a 100 μm nozzle by the SickKids UHN Flow and Mass Cytometry Facility. Around 20 000 to 50 000 gata5GFP+ cells were obtained in one FACS experiment. Single cell suspensions post FACS were loaded into the 10x machine and RNA was extracted based on Chromium Single Cell three prime Reagent Kits v2 protocol. Libraries were constructed using the 10x v2 chemistry.	Experimental Factor: age:13\|Experimental Factor: RNA interference:morpholino mediated Gata5/6 knockdown	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina HiSeq 2500	ERP122151	Illumina HiSeq 2500 sequencing; Single cell transcriptome analysis of gata5GFP labelled WT and Gata5/6 deficient cells from early gastrulation to early somitogenesis in zebrafish	ENA FIRST PUBLIC:2021 06 02\|ENA LAST UPDATE:2021 12 01	cDNA13h_2_gata56KD_S2_L002.bam	bam	7658149967.0	75823267.0	E MTAB 9193:cDNA13h 2 gata56KD S2 L002	0:101	A:2172230233;C:1568805100;G:1703587704;T:2157181071;N:56345859	101				2172230233	1568805100	1703587704	2157181071	56345859	ERX4155257	ERS4601295	ERA2667966	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	Developmental and Stem Cell Biology, Sickkids, Toronto, Canada; Genetics and Genome Biology, Sickkids, Toronto, Canada; Molecular Genetics, University of Toronto, Toronto, Canada\|European Nucleotide Archive	1	0.66565	0.08597	0.91804	0.51772	101	B	usable mapping rate	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	Canada	2020-06-04	Multi-stage	Embryo	Embryo Imprecise	All anatomical structures
10060	10060	ERR4795364	ERX4665135	ERS5281176	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 4	E MTAB 9727:Sample 4		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 4 s	Sample 4 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 4	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN4_AH2TW3BGX5_S1_R1_cat.fastq.gz	fastq	5410723202.0	71707309.0	E MTAB 9727:Sample 4	0:75.46 1:0	A:1330416803;C:531269504;G:734031980;T:2814959249;N:45666	75	0			1330416803	531269504	734031980	2814959249	45666	ERX4665135	ERS5281176	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.33614	0.21532	0.99019	0.41002	76	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10061	10061	ERR4795365	ERX4665135	ERS5281176	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 4	E MTAB 9727:Sample 4		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 4 s	Sample 4 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 4	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN4_AH2TW3BGX5_S1_R2_cat.fastq.gz	fastq	5414184547.0	71707309.0	E MTAB 9727:Sample 4 1	0:0 1:75.50	A:1582938104;C:1052565419;G:1177180395;T:1600161662;N:1338967	0	75			1582938104	1052565419	1177180395	1600161662	1338967	ERX4665135	ERS5281176	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.84976	0.28521	0.85038	0.5124	75	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10062	10062	ERR4795362	ERX4665134	ERS5281175	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 3	E MTAB 9727:Sample 3		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 3 s	Sample 3 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 3	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN3_AHY3WGBGX3_S7_R1_cat.fastq.gz	fastq	4823193891.0	63965519.0	E MTAB 9727:Sample 3	0:75.40 1:0	A:1323978162;C:446831209;G:581746343;T:2470114091;N:524086	75	0			1323978162	446831209	581746343	2470114091	524086	ERX4665134	ERS5281175	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.36549	0.19722	0.95552	0.4702	76	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10063	10063	ERR4795363	ERX4665134	ERS5281175	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 3	E MTAB 9727:Sample 3		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 3 s	Sample 3 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 3	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN3_AHY3WGBGX3_S7_R2_cat.fastq.gz	fastq	4828889391.0	63965519.0	E MTAB 9727:Sample 3 1	0:0 1:75.49	A:1434099697;C:964508763;G:893584575;T:1534727600;N:1968756	0	75			1434099697	964508763	893584575	1534727600	1968756	ERX4665134	ERS5281175	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.85811	0.26231	0.82731	0.48618	76	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10064	10064	ERR4795360	ERX4665133	ERS5281174	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 2	E MTAB 9727:Sample 2		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 2 s	Sample 2 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 2	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN2_AHY3WGBGX3_S6_R1_cat.fastq.gz	fastq	5598371393.0	74235388.0	E MTAB 9727:Sample 2	0:75.41 1:0	A:1526486973;C:558020193;G:717587491;T:2795646081;N:630655	75	0			1526486973	558020193	717587491	2795646081	630655	ERX4665133	ERS5281174	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.28059	0.19252	0.96404	0.4874	75	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10065	10065	ERR4795361	ERX4665133	ERS5281174	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 2	E MTAB 9727:Sample 2		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 2 s	Sample 2 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 2	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN2_AHY3WGBGX3_S6_R2_cat.fastq.gz	fastq	5600151371.0	74235388.0	E MTAB 9727:Sample 2 1	0:0 1:75.44	A:1850568176;C:1035622971;G:1071138281;T:1640475883;N:2346060	0	75			1850568176	1035622971	1071138281	1640475883	2346060	ERX4665133	ERS5281174	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.7679	0.30371	0.82651	0.43401	75	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10066	10066	ERR4795358	ERX4665132	ERS5281173	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 1	E MTAB 9727:Sample 1		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 1 s	Sample 1 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN1_AHY3WGBGX3_S5_R1_cat.fastq.gz	fastq	4996502316.0	66258508.0	E MTAB 9727:Sample 1	0:75.41 1:0	A:1330744174;C:451622042;G:591151500;T:2622422664;N:561936	75	0			1330744174	451622042	591151500	2622422664	561936	ERX4665132	ERS5281173	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.29754	0.17671	0.9669	0.45463	75	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10067	10067	ERR4795359	ERX4665132	ERS5281173	ERP124847	PRJEB41113	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E-MTAB-9727	Transcriptome Analysis	The mesothelium forms epithelial membranes that line the bodies cavities and surround the internal organs. Mesothelia widely contribute to organ homeostasis and regeneration and their dysregulation can result in congenital anomalies of the viscera ventral wall defects and mesothelioma tumors. Nonetheless the embryonic ontogeny and developmental regulation of mesothelium formation has remained uncharted. Here we combine genetic lineage tracing in toto live imaging and single cell transcriptomics in zebrafish to track mesothelial progenitor origins from the lateral plate mesoderm LPM. Our single cell analysis uncovers a post gastrulation gene expression signature centered on hand2 that delineates distinct progenitor populations within the forming LPM. Combining gene expression analysis and imaging of transgenic reporter zebrafish embryos we chart the origin of mesothelial progenitors to the lateral most hand2 expressing LPM and confirm evolutionary conservation in mouse. Our time lapse imaging of transgenic hand2 reporter embryos captures zebrafish mesothelium formation documenting the coordinated cell movements that form pericardium and visceral and parietal peritoneum. We establish that the primordial germ cells migrate associated with the forming mesothelium as ventral migration boundary. Functionally hand2 mutants fail to close the ventral mesothelium due to perturbed migration of mesothelium progenitors. Analyzing mouse and human mesothelioma tumors hypothesized to emerge from transformed mesothelium we find de novo expression of LPM associated transcription factors and in particular of Hand2 indicating the re initiation of a developmental transcriptional program in mesothelioma. Taken together our work outlines a genetic and developmental signature of mesothelial origins centered around Hand2 contributing to our understanding of mesothelial pathologies and mesothelioma.	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14		Protocols: Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse trans…	Sample 1	E MTAB 9727:Sample 1		isolate:not applicable\|organism:Danio rerio\|sex:mixed\|age:10\|developmental stage:tailbud stage\|organism part:lateral plate mesoderm plate\|genotype:drl:mCherry\|ENA FIRST PUBLIC:2021 04 01\|ENA LAST UPDATE:2021 09 01							NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	E MTAB 9727:Sample 1 s	Sample 1 s	Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	Wildtype stain and drl:mCherry positive zebrafish embryos were grown until tailbud stage and chorions were removed by incubating in 1 mg/mL Pronase Roche followed by washing in E3 medium. Around 300 embryos were used to generate the sample. The E3 medium was replaced by 1X PBS and the embryos were stored on ice until further processing. The embryos were dissociated using 2 mg/mL collagenase IV Worthington in DMEM high glucose 4.5g/l and NaHCO3 without xxx glutamine and sodium pyruvate Sigma Aldrich and incubated for 5 min in a water bath at 37oC. The embryonic tissues were triturated into a single cell suspension by pipetting carefully up and down. When the embryos were not yet dissociated sufficiently they were incubated for another 5 min. Cells were filtered through a 35 μm cell strainer Falcon round bottom tubes with cell strainer cap and centrifuged at 6000 rpm for 30 sec. Cell pellets were resuspended in 1X HBSS Gibco containing 2% FBS and subjected to an additional round of centrifugation and resuspension. post washing the cells were resuspended in 1X PBS. drl:mCherry positive cells were sorted using a FACS Aria III cell sorter BD Bioscience. Cells were gated based on size and forward scattering to exclude debris and doublets. The gating for the negative population was determined based on wildtype tailbud staged embryos. See also Figure S2 for gate setting. The SORTseq single cell RNA sequencing protocol was carried out as described previously Muraro et al. 2016. Live mCherry positive single cells were sorted in four 384 well plates BioRad containing 5 μl of CEL Seq2 primer solution in mineral oil 24 bp polyT stretch a 4 bp random molecular barcode UMI a cell specific barcode the 5′Illumina TruSeq small RNA kit adaptor and a T7 promoter provided by Single Cell Discoveries. post sorting the plates were immediately placed on ice and stored at −80°C. In brief ERCC Spike in RNA 0.02 μL of 1:50000 dilution was added to each well before cell lysis with heat shocking. Reverse transcription an…	Experimental Factor: replicate:lateral plate mesoderm plate 1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	RANDOM	SINGLE	ILLUMINA	NextSeq 500	ERP124847	NextSeq 500 sequencing; Hand2 delineates mesothelium progenitors and is reactivated in mesothelioma	ENA FIRST PUBLIC:2022 01 14\|ENA LAST UPDATE:2022 01 14	SN1_AHY3WGBGX3_S5_R2_cat.fastq.gz	fastq	4999908389.0	66258508.0	E MTAB 9727:Sample 1 1	0:0 1:75.46	A:1591945117;C:933968124;G:986108614;T:1485824022;N:2062512	0	75			1591945117	933968124	986108614	1485824022	2062512	ERX4665132	ERS5281173	ERA3048543	University of Zurich\|European Nucleotide Archive	University of Zurich\|European Nucleotide Archive	1	0.81333	0.2466	0.83027	0.51572	75	B	usable mapping rate	illumina	nextseq	unknown	random_priming	trueseq	sc	single_cell_plate	celseq	Switzerland	2021-04-01	Gastrula	Embryo	Embryo Imprecise	All anatomical structures
10186	10186	ERR6212423	ERX5847531	ERS7094943	ERP130388	PRJEB46176	scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	E-MTAB-10379_3	Transcriptome Analysis	To analyse lesion induced gene regulation in macrophage and microglia at single cell resolution we performed single cell RNAseq on FACS isolated mpeg1.1:GFP cells from the spinal cord at 24 hours post lesion hpl post spinal injury at 3 dpf dpf compared to age matched uninjured animals.	ENA FIRST PUBLIC:2021 07 08\|ENA LAST UPDATE:2021 07 08		Protocols: Trunks containing the lesion sites or equivalent site from unlesioned fish are collected and kept in PBS on ice.Incubate the trunks up to 300 in 1 mL of 1X Trypsin EDTA at 37C for 5 7 mins.Stop dissociation by adding FBS to a final concentration of 5%.Centrifuge at 200g for 7 mins.Discard Sups.Resuspend in 500 uL of PBS.Add the cell suspension into a 40 uM cell strainer.Centrifuge at 200g for 7 mins.Resuspend in the buffer for FACS PBS 5% FBS Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines	Naive	SAMEA9361876	University Of Edinburgh	ENA first public:2021 07 08\|ENA last update:2021 07 08\|External Id:SAMEA9361876\|INSDC center alias:UOE\|INSDC center name:University Of Edinburgh\|INSDC first public:2021 07 08T11:18:57Z\|INSDC last update:2021 07 08T11:18:57Z\|INSDC status:public\|Submitter Id:E MTAB 10379 3:Naive\|age:4\|broker name:ArrayExpress\|cell type:mixed macrophage and microglia\|common name:zebrafish\|developmental stage:larval day 4\|immunophenotype:mpeg1.1 positive\|organism part:spinal cord\|sample name:E MTAB 10379 3:Naive\|sex:mixed\|strain:WIK							Illumina NovaSeq 6000 sequencing; scRNAseq of mpeg1.1+ cells from lesi1d and unlesi1d zebrafish larvae spinal cord	E MTAB 10379 3:Naive p	Naive p	scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	Trunks containing the lesion sites or equivalent site from unlesioned fish are collected and kept in PBS on ice.Incubate the trunks up to 300 in 1 mL of 1X Trypsin EDTA at 37C for 5 7 mins.Stop dissociation by adding FBS to a final concentration of 5%.Centrifuge at 200g for 7 mins.Discard Sups.Resuspend in 500 uL of PBS.Add the cell suspension into a 40 uM cell strainer.Centrifuge at 200g for 7 mins.Resuspend in the buffer for FACS PBS 5% FBS Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines	Experimental Factor: injury:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	Oligo-dT	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP130388	Illumina NovaSeq 6000 sequencing; scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	ENA FIRST PUBLIC:2021 07 08\|ENA LAST UPDATE:2021 07 08	Mpeg_Naive_I1.fastq.gz Mpeg_Naive_R1.fastq.gz Mpeg_Naive_R2.fastq.gz	fastq fastq fastq	60168324125.0	481346593.0	E MTAB 10379 3:Naive	0:8 1:27 2:90	A:12424400694;C:9366563781;G:10029312355;T:11497237299;N:3679241	8	27	90		12424400694	9366563781	10029312355	11497237299	3679241	ERX5847531	ERS7094943	ERA5186263	University Of Edinburgh\|European Nucleotide Archive	University Of Edinburgh\|European Nucleotide Archive	1	0.94834	0.09482	0.802	0.5277	90	B	usable mapping rate	illumina	novaseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	United Kingdom	2021-07-08	Larval	Larval	Multi-tissue	Multi-system
10187	10187	ERR6212422	ERX5847530	ERS7094942	ERP130388	PRJEB46176	scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	E-MTAB-10379_3	Transcriptome Analysis	To analyse lesion induced gene regulation in macrophage and microglia at single cell resolution we performed single cell RNAseq on FACS isolated mpeg1.1:GFP cells from the spinal cord at 24 hours post lesion hpl post spinal injury at 3 dpf dpf compared to age matched uninjured animals.	ENA FIRST PUBLIC:2021 07 08\|ENA LAST UPDATE:2021 07 08		Protocols: Trunks containing the lesion sites or equivalent site from unlesioned fish are collected and kept in PBS on ice.Incubate the trunks up to 300 in 1 mL of 1X Trypsin EDTA at 37C for 5 7 mins.Stop dissociation by adding FBS to a final concentration of 5%.Centrifuge at 200g for 7 mins.Discard Sups.Resuspend in 500 uL of PBS.Add the cell suspension into a 40 uM cell strainer.Centrifuge at 200g for 7 mins.Resuspend in the buffer for FACS PBS 5% FBS Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines	Lesi1d	SAMEA9361875	University Of Edinburgh	ENA first public:2021 07 08\|ENA last update:2021 07 08\|External Id:SAMEA9361875\|INSDC center alias:UOE\|INSDC center name:University Of Edinburgh\|INSDC first public:2021 07 08T11:18:57Z\|INSDC last update:2021 07 08T11:18:57Z\|INSDC status:public\|Submitter Id:E MTAB 10379 3:Lesi1d\|age:4\|broker name:ArrayExpress\|cell type:mixed macrophage and microglia\|common name:zebrafish\|developmental stage:larval day 4\|immunophenotype:mpeg1.1 positive\|injury:spinal cord lesion\|organism part:spinal cord\|sample name:E MTAB 10379 3:Lesi1d\|sex:mixed\|strain:WIK							Illumina NovaSeq 6000 sequencing; scRNAseq of mpeg1.1+ cells from lesi1d and unlesi1d zebrafish larvae spinal cord	E MTAB 10379 3:Lesioned p	Lesioned p	scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	Trunks containing the lesion sites or equivalent site from unlesioned fish are collected and kept in PBS on ice.Incubate the trunks up to 300 in 1 mL of 1X Trypsin EDTA at 37C for 5 7 mins.Stop dissociation by adding FBS to a final concentration of 5%.Centrifuge at 200g for 7 mins.Discard Sups.Resuspend in 500 uL of PBS.Add the cell suspension into a 40 uM cell strainer.Centrifuge at 200g for 7 mins.Resuspend in the buffer for FACS PBS 5% FBS Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines Following cell dissociation and FAC sorting samples were processed on the 10X Chromium platform using 10X Single Cell three prime v3 chemistry following the manufacturer's guidelines	Experimental Factor: injury:spinal cord lesion	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	Oligo-dT	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP130388	Illumina NovaSeq 6000 sequencing; scRNAseq of mpeg1.1+ cells from lesioned and unlesioned zebrafish larvae spinal cord	ENA FIRST PUBLIC:2021 07 08\|ENA LAST UPDATE:2021 07 08	Mpeg_Lesioned_I1.fastq.gz Mpeg_Lesioned_R1.fastq.gz Mpeg_Lesioned_R2.fastq.gz	fastq fastq fastq	61701873000.0	493614984.0	E MTAB 10379 3:Lesioned	0:8 1:27 2:90	A:12952533019;C:9222397450;G:9920185920;T:12326470894;N:3761277	8	27	90		12952533019	9222397450	9920185920	12326470894	3761277	ERX5847530	ERS7094942	ERA5186263	University Of Edinburgh\|European Nucleotide Archive	University Of Edinburgh\|European Nucleotide Archive	1	0.93082	0.10862	0.80302	0.54924	90	B	usable mapping rate	illumina	novaseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	United Kingdom	2021-07-08	Larval	Larval	Multi-tissue	Multi-system
11017	11017	ERR9750937	ERX9298876	ERS12023662	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 2 Uninjured	SAMEA14418104	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418104\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 2 Uninjured\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|organism part:heart\|sample name:E MTAB 10643:Expt 2 Uninjured\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 2 Uninjured p	Expt 2 Uninjured p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:n1\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E2_Uninjured_S2_L002_I1_001.fastq.gz E2_Uninjured_S2_L002_I2_001.fastq.gz E2_Uninjured_S2_L002_R1_001.fastq.gz E2_Uninjured_S2_L002_R2_001.fastq.gz	fastq fastq fastq fastq	9325673775.0	69079065.0	E MTAB 10643:Expt 2 Uninjured	0:10 1:10 2:28 3:87	A:1619571910;C:1420805235;G:1471531128;T:1496998088;N:972294	10	10	28	87	1619571910	1420805235	1471531128	1496998088	972294	ERX9298876	ERS12023662	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.93798	0.11364	0.84843	0.53531	87	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11018	11018	ERR9750936	ERX9298875	ERS12023661	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 2 Sham	SAMEA14418103	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418103\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 2 Sham\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:sham surgery\|organism part:heart\|sample name:E MTAB 10643:Expt 2 Sham\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 2 Sham p	Expt 2 Sham p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:sham surgery\|Experimental Factor: time:3\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E2_Sham_S1_L002_I1_001.fastq.gz E2_Sham_S1_L002_I2_001.fastq.gz E2_Sham_S1_L002_R1_001.fastq.gz E2_Sham_S1_L002_R2_001.fastq.gz	fastq fastq fastq fastq	11328674040.0	83916104.0	E MTAB 10643:Expt 2 Sham	0:10 1:10 2:28 3:87	A:1955327720;C:1746332187;G:1777122701;T:1820705338;N:1213102	10	10	28	87	1955327720	1746332187	1777122701	1820705338	1213102	ERX9298875	ERS12023661	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.92847	0.09981	0.85707	0.46947	87	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11019	11019	ERR9750935	ERX9298874	ERS12023660	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 2 3d post amputation	SAMEA14418102	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418102\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 2 3d post amputation\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:20 percent removal of the cardiac ventricle apex\|organism part:heart\|sample name:E MTAB 10643:Expt 2 3d post amputation\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 2 3d post amputation p	Expt 2 3d post amputation p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:20 percent removal of the cardiac ventricle apex\|Experimental Factor: time:3\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E2_3dpa_S3_L002_I1_001.fastq.gz E2_3dpa_S3_L002_I2_001.fastq.gz E2_3dpa_S3_L002_R1_001.fastq.gz E2_3dpa_S3_L002_R2_001.fastq.gz	fastq fastq fastq fastq	16111486440.0	119344344.0	E MTAB 10643:Expt 2 3d post amputation	0:10 1:10 2:28 3:87	A:2610144058;C:2617445332;G:2496763759;T:2656876552;N:1728227	10	10	28	87	2610144058	2617445332	2496763759	2656876552	1728227	ERX9298874	ERS12023660	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.94868	0.07071	0.89221	0.54333	87	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11020	11020	ERR9750934	ERX9298873	ERS12023659	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 3 3d post amputation + MMP inhibitor	SAMEA14418101	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418101\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 3 3d post amputation + MMP inhibitor\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:20 percent removal of the cardiac ventricle apex\|organism part:heart\|sample name:E MTAB 10643:Expt 3 3d post amputation + MMP inhibitor\|sex:male\|strain:AB\|treatment:matrix metalloproteinase inhibitor							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 3 3d post amputation + MMP inhibitor p	Expt 3 3d post amputation + MMP inhibitor p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Zebrafish were treated with MMP inhibitor NSC405020 for three days following surgery. Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:20 percent removal of the cardiac ventricle apex\|Experimental Factor: time:3\|Experimental Factor: treatment:matrix metalloproteinase inhibitor	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E2_3dpa_inhibitor_S4_L002_I1_001.fastq.gz E2_3dpa_inhibitor_S4_L002_I2_001.fastq.gz E2_3dpa_inhibitor_S4_L002_R1_001.fastq.gz E2_3dpa_inhibitor_S4_L002_R2_001.fastq.gz	fastq fastq fastq fastq	8621019270.0	63859402.0	E MTAB 10643:Expt 3 3d post amputation + MMP inhibitor	0:10 1:10 2:28 3:87	A:1438763050;C:1365497247;G:1350808021;T:1399773154;N:926502	10	10	28	87	1438763050	1365497247	1350808021	1399773154	926502	ERX9298873	ERS12023659	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.92796	0.08671	0.87176	0.55088	87	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11021	11021	ERR9750933	ERX9298872	ERS12023658	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 1 Uninjured	SAMEA14418100	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418100\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 1 Uninjured\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|organism part:heart\|sample name:E MTAB 10643:Expt 1 Uninjured\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 1 Uninjured p	Expt 1 Uninjured p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:n1\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E1_Uninjured_S1_L002_I1_001.fastq.gz E1_Uninjured_S1_L002_R1_001.fastq.gz E1_Uninjured_S1_L002_R2_001.fastq.gz	fastq fastq fastq	8575585211.0	67524293.0	E MTAB 10643:Expt 1 Uninjured	0:8 1:28 2:91	A:1801503847;C:1312196506;G:1367890726;T:1663053783;N:65801	8	28	91		1801503847	1312196506	1367890726	1663053783	65801	ERX9298872	ERS12023658	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.9169	0.11202	0.81576	0.54253	91	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11022	11022	ERR9750932	ERX9298871	ERS12023657	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 1 7d post amputation	SAMEA14418099	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418099\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 1 7d post amputation\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:20 percent removal of the cardiac ventricle apex\|organism part:heart\|sample name:E MTAB 10643:Expt 1 7d post amputation\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 1 7d post amputation p	Expt 1 7d post amputation p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:20 percent removal of the cardiac ventricle apex\|Experimental Factor: time:7\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E1_7d_S3_L002_I1_001.fastq.gz E1_7d_S3_L002_R1_001.fastq.gz E1_7d_S3_L002_R2_001.fastq.gz	fastq fastq fastq	17341796660.0	136549580.0	E MTAB 10643:Expt 1 7d post amputation	0:8 1:28 2:91	A:3506600812;C:2774617588;G:2814960106;T:3329701112;N:132162	8	28	91		3506600812	2774617588	2814960106	3329701112	132162	ERX9298871	ERS12023657	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.90616	0.09608	0.82446	0.46632	91	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11023	11023	ERR9750931	ERX9298870	ERS12023656	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 1 3d post amputation	SAMEA14418098	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418098\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 1 3d post amputation\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:20 percent removal of the cardiac ventricle apex\|organism part:heart\|sample name:E MTAB 10643:Expt 1 3d post amputation\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 1 3d post amputation p	Expt 1 3d post amputation p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:20 percent removal of the cardiac ventricle apex\|Experimental Factor: time:3\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E1_3d_S2_L002_I1_001.fastq.gz E1_3d_S2_L002_R1_001.fastq.gz E1_3d_S2_L002_R2_001.fastq.gz	fastq fastq fastq	8183008018.0	64433134.0	E MTAB 10643:Expt 1 3d post amputation	0:8 1:28 2:91	A:1630460899;C:1330248096;G:1326144211;T:1576499358;N:62630	8	28	91		1630460899	1330248096	1326144211	1576499358	62630	ERX9298870	ERS12023656	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.88759	0.08679	0.82948	0.47863	91	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
11024	11024	ERR9750930	ERX9298869	ERS12023655	ERP137743	PRJEB52989	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E-MTAB-10643	Transcriptome Analysis	Adult zebrafish hearts have the ability to regenerate. The roles of non myocytes in this process have remained elusive. Here we have performed 2 scRNAseq experiments on interstitial cells. Experiment 1 E1 included interstitial cells obtained from uninjured regenerating 3 days 7 days and 14 days post apical amputation. Experiment 2 E2 included cells from uninjured sham operated abdomen opened and regenerating 3 dy post amputation with and without xxx inhibitor NSC40520 treatment. Cells were obtained by heart dissection followed by enzymatic dissociation and FACS sorting of single viable nucleated cells.	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20		Protocols: Zebrafish were euthanized in tricaine and hearts were extracted in PBS Hearts were pooled and pre digested for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum. The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Expt 1 14d post amputation	SAMEA14418097	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.	ENA first public:2022 05 20\|ENA last update:2022 05 20\|External Id:SAMEA14418097\|INSDC center alias:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC center name:IGF CNRS INSERM Univ. Montpellier LabEx ICST F 34094 Montpellier France.\|INSDC first public:2022 05 20T16:17:49Z\|INSDC last update:2022 05 20T16:17:49Z\|INSDC status:public\|Submitter Id:E MTAB 10643:Expt 1 14d post amputation\|age:6\|broker name:ArrayExpress\|cell type:interstitial cell\|common name:zebrafish\|developmental stage:adult\|genotype:wild type genotype\|injury:20 percent removal of the cardiac ventricle apex\|organism part:heart\|sample name:E MTAB 10643:Expt 1 14d post amputation\|sex:male\|strain:AB							Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	E MTAB 10643:Expt 1 14d post amputation p	Expt 1 14d post amputation p	scRNAseq of post amputation regenerating zebrafish cardiac ventricle	Zebrafish were euthanized in tricaine and hearts were extracted in PBS "Hearts were pooled and ""pre digested"" for 1h in 1/10 TrypLE/HBSS No CaMg on ice. Hearts were then digested in Collagenase IV 5mg/ml + Collagenase II 5mg/ml in HBSS with 12.5uM CaCl2 at 32 deg on a shaker 800rpm for 45 min. Following this tissue was gently dissociated and passed through a cell strainer 40um spun down 5min 300g and resuspended in HBSS 2% fetal calf serum." The heart was exposed and approximately 20% of the ventricle removed Amplified cDNA was used for three prime RNA seq library generation RNA seq libraries were prepared following the manufacturer's user guide 10x Genomics.	Experimental Factor: injury:20 percent removal of the cardiac ventricle apex\|Experimental Factor: time:14\|Experimental Factor: treatment:n1	RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	PolyA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP137743	Illumina NovaSeq 6000 sequencing; scRNAseq of post amputation regenerating zebrafish cardiac ventricle	ENA FIRST PUBLIC:2022 05 20\|ENA LAST UPDATE:2022 05 20	E1_14d_S4_L002_I1_001.fastq.gz E1_14d_S4_L002_R1_001.fastq.gz E1_14d_S4_L002_R2_001.fastq.gz	fastq fastq fastq	12911908390.0	101668570.0	E MTAB 10643:Expt 1 14d post amputation	0:8 1:28 2:91	A:2700371496;C:1971281481;G:2062280628;T:2517806408;N:99857	8	28	91		2700371496	1971281481	2062280628	2517806408	99857	ERX9298869	ERS12023655	ERA14538536	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	IGF, CNRS, INSERM, Univ. Montpellier, LabEx ICST, F-34094 Montpellier, France.\|European Nucleotide Archive	1	0.90771	0.11048	0.81523	0.53024	91	B	usable mapping rate	illumina	novaseq_era	3prime	poly_a	unknown	sc	single_cell_droplet	10x	France	2022-05-20	Adult	Adult	Heart	Cardiovascular System
19476	19476	ERR14208827	ERX13611047	ERS22979745	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	Scrambled 1 sample	SAMEA117628607	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 004\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 004\|scientific name:Danio rerio							Raw reads: Scrambled 1 sample	webin reads Scrambled 1 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: Scrambled 1 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F04.bam	bam	8497741.0	104283.0	webin reads Scrambled 1 sample	0:81.49	A:2628864;C:1112827;G:2146146;T:2609768;N:136	81				2628864	1112827	2146146	2609768	136	ERX13611047	ERS22979745	ERA31123309	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19477	19477	ERR14208813	ERX13611033	ERS22979748	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	ythdf2KO 1 sample	SAMEA117628610	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 007\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 007\|scientific name:Danio rerio							Raw reads: ythdf2KO 1 sample	webin reads ythdf2KO 1 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: ythdf2KO 1 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	D09.bam	bam	673213184.0	8122011.0	webin reads ythdf2KO 1 sample	0:82.89	A:180836688;C:107076381;G:193711319;T:191586188;N:2608	82				180836688	107076381	193711319	191586188	2608	ERX13611033	ERS22979748	ERA31123295	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19478	19478	ERR14208811	ERX13611031	ERS22979755	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 6 sample	SAMEA117628617	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 014\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 014\|scientific name:Danio rerio							Raw reads: GFP 6 sample	webin reads GFP 6 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 6 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	H01.bam	bam	337553787.0	3153658.0	webin reads GFP 6 sample	0:107.04	A:81816649;C:55524538;G:113686171;T:86525390;N:1039	107				81816649	55524538	113686171	86525390	1039	ERX13611031	ERS22979755	ERA31123293	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19479	19479	ERR14208821	ERX13611041	ERS22979752	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	ythdf2KO 5 sample	SAMEA117628614	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 011\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 011\|scientific name:Danio rerio							Raw reads: ythdf2KO 5 sample	webin reads ythdf2KO 5 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: ythdf2KO 5 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	G10.bam	bam	226771232.0	2050630.0	webin reads ythdf2KO 5 sample	0:110.59	A:56944731;C:30766256;G:83820146;T:55239221;N:878	110				56944731	30766256	83820146	55239221	878	ERX13611041	ERS22979752	ERA31123303	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19480	19480	ERR14208818	ERX13611038	ERS22979750	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	ythdf2KO 3 sample	SAMEA117628612	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 009\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 009\|scientific name:Danio rerio							Raw reads: ythdf2KO 3 sample	webin reads ythdf2KO 3 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: ythdf2KO 3 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	G08.bam	bam	351425328.0	3771763.0	webin reads ythdf2KO 3 sample	0:93.17	A:83933175;C:50138103;G:128300798;T:89051913;N:1339	93				83933175	50138103	128300798	89051913	1339	ERX13611038	ERS22979750	ERA31123300	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19481	19481	ERR14208826	ERX13611046	ERS22979758	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	YTHDF2 3 sample	SAMEA117628620	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 017\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 017\|scientific name:Danio rerio							Raw reads: YTHDF2 3 sample	webin reads YTHDF2 3 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: YTHDF2 3 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	E01.bam	bam	181017083.0	1670383.0	webin reads YTHDF2 3 sample	0:108.37	A:38910474;C:31607688;G:67095095;T:43403227;N:599	108				38910474	31607688	67095095	43403227	599	ERX13611046	ERS22979758	ERA31123308	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19482	19482	ERR14208815	ERX13611035	ERS22979749	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	ythdf2KO 2 sample	SAMEA117628611	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 008\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 008\|scientific name:Danio rerio							Raw reads: ythdf2KO 2 sample	webin reads ythdf2KO 2 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: ythdf2KO 2 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	D10.bam	bam	282414832.0	2675188.0	webin reads ythdf2KO 2 sample	0:105.57	A:73651155;C:41694223;G:96485766;T:70582706;N:982	105				73651155	41694223	96485766	70582706	982	ERX13611035	ERS22979749	ERA31123297	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19483	19483	ERR14208804	ERX13611024	ERS22979742	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 1 sample	SAMEA117628604	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 001\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 001\|scientific name:Danio rerio							Raw reads: GFP 1 sample	webin reads GFP 1 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 1 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F01.bam	bam	8036009.0	97716.0	webin reads GFP 1 sample	0:82.24	A:2445867;C:1074319;G:2086526;T:2429153;N:144	82				2445867	1074319	2086526	2429153	144	ERX13611024	ERS22979742	ERA31123286	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19484	19484	ERR14208829	ERX13611049	ERS22979747	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	Scrambled 3 sample	SAMEA117628609	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 006\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 006\|scientific name:Danio rerio							Raw reads: Scrambled 3 sample	webin reads Scrambled 3 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: Scrambled 3 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F06.bam	bam	90051514.0	910768.0	webin reads Scrambled 3 sample	0:98.87	A:30132090;C:13700682;G:16323331;T:29894332;N:1079	98				30132090	13700682	16323331	29894332	1079	ERX13611049	ERS22979747	ERA31123311	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19485	19485	ERR14208809	ERX13611029	ERS22979753	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 4 sample	SAMEA117628615	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 012\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 012\|scientific name:Danio rerio							Raw reads: GFP 4 sample	webin reads GFP 4 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 4 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	G11.bam	bam	72278736.0	751207.0	webin reads GFP 4 sample	0:96.22	A:17589630;C:11153310;G:25212154;T:18323416;N:226	96				17589630	11153310	25212154	18323416	226	ERX13611029	ERS22979753	ERA31123291	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19486	19486	ERR14208806	ERX13611026	ERS22979743	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 2 sample	SAMEA117628605	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 002\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 002\|scientific name:Danio rerio							Raw reads: GFP 2 sample	webin reads GFP 2 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 2 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F02.bam	bam	19196592.0	230075.0	webin reads GFP 2 sample	0:83.44	A:5957309;C:2709511;G:4434875;T:6094552;N:345	83				5957309	2709511	4434875	6094552	345	ERX13611026	ERS22979743	ERA31123288	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19487	19487	ERR14208810	ERX13611030	ERS22979754	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 5 sample	SAMEA117628616	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 013\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 013\|scientific name:Danio rerio							Raw reads: GFP 5 sample	webin reads GFP 5 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 5 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	G12.bam	bam	431708790.0	5173015.0	webin reads GFP 5 sample	0:83.45	A:119077902;C:68329173;G:126879243;T:117420803;N:1669	83				119077902	68329173	126879243	117420803	1669	ERX13611030	ERS22979754	ERA31123292	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19488	19488	ERR14208828	ERX13611048	ERS22979746	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	Scrambled 2 sample	SAMEA117628608	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 005\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 005\|scientific name:Danio rerio							Raw reads: Scrambled 2 sample	webin reads Scrambled 2 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: Scrambled 2 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F05.bam	bam	32695808.0	357320.0	webin reads Scrambled 2 sample	0:91.50	A:10989624;C:4719485;G:5954777;T:11031603;N:319	91				10989624	4719485	5954777	11031603	319	ERX13611048	ERS22979746	ERA31123310	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19489	19489	ERR14208819	ERX13611039	ERS22979751	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	ythdf2KO 4 sample	SAMEA117628613	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 010\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 010\|scientific name:Danio rerio							Raw reads: ythdf2KO 4 sample	webin reads ythdf2KO 4 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: ythdf2KO 4 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	G09.bam	bam	254231539.0	2766205.0	webin reads ythdf2KO 4 sample	0:91.91	A:63984512;C:40328422;G:86431821;T:63485754;N:1030	91				63984512	40328422	86431821	63485754	1030	ERX13611039	ERS22979751	ERA31123301	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19490	19490	ERR14208808	ERX13611028	ERS22979744	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	GFP 3 sample	SAMEA117628606	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 003\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 003\|scientific name:Danio rerio							Raw reads: GFP 3 sample	webin reads GFP 3 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: GFP 3 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	F03.bam	bam	35932988.0	419944.0	webin reads GFP 3 sample	0:85.57	A:11252929;C:5321445;G:8032297;T:11325814;N:503	85				11252929	5321445	8032297	11325814	503	ERX13611028	ERS22979744	ERA31123290	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19491	19491	ERR14208825	ERX13611045	ERS22979757	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	YTHDF2 2 sample	SAMEA117628619	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 016\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 016\|scientific name:Danio rerio							Raw reads: YTHDF2 2 sample	webin reads YTHDF2 2 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: YTHDF2 2 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	D12.bam	bam	626993184.0	7599502.0	webin reads YTHDF2 2 sample	0:82.50	A:182701108;C:91008174;G:172822143;T:180459253;N:2506	82				182701108	91008174	172822143	180459253	2506	ERX13611045	ERS22979757	ERA31123307	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
19492	19492	ERR14208822	ERX13611042	ERS22979756	ERP167299	PRJEB83709	YTHDF2 and ASD DM	3296f78e-e1c0-43ea-968c-53fe04e67615	Other	Among autistic individuals a subphenotype of disproportionate megalencephaly ASD DM seen at three years of age is associated with co occurring intellectual disability and poorer prognoses later in life. However many of the genes contributing to ASD DM have yet to be delineated. In this study we identified additional ASD DM candidate genes with the aim to better define the genetic etiology of this subphenotype of autism. We expanded the previously studied sample size of ASD DM individuals ten fold by including probands from the Autism Phenome Project and Simons Simplex Collection totaling 766 autistic individuals meeting the criteria for megalencephaly or macrocephaly and revealing 154 candidate ASD DM genes harboring de novo protein impacting variants. Our findings include fourteen high confidence autism genes and seven genes previously associated with DM. Five impacted genes have previously been associated with both autism and DM including CHD8 and PTEN. By performing functional network analysis we expanded to additional candidate genes including one previously implicated in ASD DM PIK3CA as well as 184 additional genes previously implicated in ASD or DM alone. Using zebrafish we modeled a de novo tandem duplication impacting YTHDF2 encoding an N6 methyladenosine m6A mRNA reader in an ASD DM proband. Testing zebrafish CRISPR knockdown led to reduced head/brain size while overexpressing YTHDF2 resulted in increased head and brain size matching that of the proband. Single cell transcriptomes of YTHDF2 gain of function larvae point to reduced expression of Fragile X syndrome associated FMRP target genes globally and in the developing brain providing insight into the mechanism underlying autistic phenotypes. We additionally discovered a variant impacting a different gene encoding an m6A reader YTHDC1 in our ASD DM cohort. Though we highlight only two cases to date our study provides support for the m6A RNA modification pathway as potentially contributing to this severe form of autism.	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01		Single cell RNA sequencing of zebrafish heads	YTHDF2 1 sample	SAMEA117628618	UNIVERSITY OF CALIFORNIA - DAVIS	ENA first public:2025 02 01\|INSDC center name:UNIVERSITY OF CALIFORNIA DAVIS\|INSDC status:public\|Submitter Id:Sample 015\|collection date:2022 08 07\|common name:zebrafish\|dev stage:72 hpf\|geographic location country and/or sea:USA\|sample name:Sample 015\|scientific name:Danio rerio							Raw reads: YTHDF2 1 sample	webin reads YTHDF2 1 sample		unspecified			RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina NovaSeq 6000	ERP167299	Raw reads: YTHDF2 1 sample	ENA FIRST PUBLIC:2025 02 01\|ENA LAST UPDATE:2025 02 01	D11.bam	bam	120748751.0	1368887.0	webin reads YTHDF2 1 sample	0:88.21	A:34957526;C:19985010;G:32927086;T:32878659;N:470	88				34957526	19985010	32927086	32878659	470	ERX13611042	ERS22979756	ERA31123304	UNIVERSITY OF CALIFORNIA - DAVIS\|European Nucleotide Archive	UNIVERSITY OF CALIFORNIA - DAVIS							B	usable mapping rate	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_generic	generic-scrnaseq-only	United States	2025-02-01	Larval	Larval	Head	Nervous System
25107	25107	SRR25605432	SRX21332628	SRS18578260	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep3	GSM7702835		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702835	GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq	GSM7702835 r1	GSM7702835	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_GFP_pos_S4_L001_R2_001.fastq.gz 2707_GFP_pos_S4_L001_R1_001.fastq.gz 2707_GFP_pos_S4_L001_I2_001.fastq.gz 2707_GFP_pos_S4_L001_I1_001.fastq.gz	fastq fastq fastq fastq	21603309774.0	156545723.0	GSM7702835 r1	0:10 1:10 2:28 3:90	A:3960451458;C:3105338977;G:3480072824;T:3541218595;N:2033216	10	10	28	90	3960451458	3105338977	3480072824	3541218595	2033216	SRX21332628	SRS18578260	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94286	0.14175	0.78395	0.51541	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25108	25108	SRR25605433	SRX21332628	SRS18578260	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep3	GSM7702835		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702835	GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq	GSM7702835 r1	GSM7702835	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_GFP_pos_S4_L002_I1_001.fastq.gz 2707_GFP_pos_S4_L002_I2_001.fastq.gz 2707_GFP_pos_S4_L002_R1_001.fastq.gz 2707_GFP_pos_S4_L002_R2_001.fastq.gz	fastq fastq fastq fastq	22413465270.0	162416415.0	GSM7702835 r2	0:10 1:10 2:28 3:90	A:4104405257;C:3220235633;G:3625448968;T:3667329471;N:58021	10	10	28	90	4104405257	3220235633	3625448968	3667329471	58021	SRX21332628	SRS18578260	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94239	0.14234	0.78338	0.50616	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25109	25109	SRR25822232	SRX21332628	SRS18578260	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep3	GSM7702835		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702835	GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq	GSM7702835 r1	GSM7702835	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_GFP_pos_S4_L001_R2_002.fastq.gz 2707_GFP_pos_S4_L001_R1_002.fastq.gz 2707_GFP_pos_S4_L001_I2_002.fastq.gz 2707_GFP_pos_S4_L001_I1_002.fastq.gz	fastq fastq fastq fastq	21424338264.0	155248828.0	GSM7702835 r3	0:10 1:10 2:28 3:90	A:3928077020;C:3077672525;G:3451667186;T:3513734857;N:1242932	10	10	28	90	3928077020	3077672525	3451667186	3513734857	1242932	SRX21332628	SRS18578260	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.9433	0.14242	0.78328	0.52384	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25110	25110	SRR25822233	SRX21332628	SRS18578260	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep3	GSM7702835		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702835	GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq	GSM7702835 r1	GSM7702835	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_GFP_pos_S4_L002_I1_002.fastq.gz 2707_GFP_pos_S4_L002_I2_002.fastq.gz 2707_GFP_pos_S4_L002_R1_002.fastq.gz 2707_GFP_pos_S4_L002_R2_002.fastq.gz	fastq fastq fastq fastq	22542479952.0	163351304.0	GSM7702835 r4	0:10 1:10 2:28 3:90	A:4130174287;C:3235362228;G:3643457599;T:3691295761;N:1327485	10	10	28	90	4130174287	3235362228	3643457599	3691295761	1327485	SRX21332628	SRS18578260	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94183	0.14124	0.78301	0.50517	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25111	25111	SRR25605434	SRX21332627	SRS18578259	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep3	GSM7702834		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702834	GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq	GSM7702834 r1	GSM7702834	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_pos_Mcherry_S3_L001_R2_001.fastq.gz 2707_pos_Mcherry_S3_L001_R1_001.fastq.gz 2707_pos_Mcherry_S3_L001_I2_001.fastq.gz 2707_pos_Mcherry_S3_L001_I1_001.fastq.gz	fastq fastq fastq fastq	20161522758.0	146097991.0	GSM7702834 r1	0:10 1:10 2:28 3:90	A:3649819934;C:2958286158;G:3267504300;T:3271384686;N:1824112	10	10	28	90	3649819934	2958286158	3267504300	3271384686	1824112	SRX21332627	SRS18578259	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94851	0.1483	0.80012	0.52549	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25112	25112	SRR25605435	SRX21332627	SRS18578259	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep3	GSM7702834		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702834	GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq	GSM7702834 r1	GSM7702834	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_pos_Mcherry_S3_L002_I1_001.fastq.gz 2707_pos_Mcherry_S3_L002_I2_001.fastq.gz 2707_pos_Mcherry_S3_L002_R1_001.fastq.gz 2707_pos_Mcherry_S3_L002_R2_001.fastq.gz	fastq fastq fastq fastq	20708273928.0	150059956.0	GSM7702834 r2	0:10 1:10 2:28 3:90	A:3746088654;C:3037032642;G:3368938151;T:3353289242;N:47351	10	10	28	90	3746088654	3037032642	3368938151	3353289242	47351	SRX21332627	SRS18578259	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94778	0.14581	0.80028	0.52885	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25113	25113	SRR25822230	SRX21332627	SRS18578259	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep3	GSM7702834		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702834	GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq	GSM7702834 r1	GSM7702834	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_pos_Mcherry_S3_L001_I1_002.fastq.gz 2707_pos_Mcherry_S3_L001_I2_002.fastq.gz 2707_pos_Mcherry_S3_L001_R1_002.fastq.gz 2707_pos_Mcherry_S3_L001_R2_002.fastq.gz	fastq fastq fastq fastq	20078121078.0	145493631.0	GSM7702834 r3	0:10 1:10 2:28 3:90	A:3634984131;C:2943891505;G:3255149272;T:3259243920;N:1157962	10	10	28	90	3634984131	2943891505	3255149272	3259243920	1157962	SRX21332627	SRS18578259	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94884	0.14622	0.80099	0.52572	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25114	25114	SRR25822231	SRX21332627	SRS18578259	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep3	GSM7702834		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep3	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702834	GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq	GSM7702834 r1	GSM7702834	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	2707_pos_Mcherry_S3_L002_I1_002.fastq.gz 2707_pos_Mcherry_S3_L002_I2_002.fastq.gz 2707_pos_Mcherry_S3_L002_R1_002.fastq.gz 2707_pos_Mcherry_S3_L002_R2_002.fastq.gz	fastq fastq fastq fastq	20884392978.0	151336181.0	GSM7702834 r4	0:10 1:10 2:28 3:90	A:3779370721;C:3059502508;G:3396295214;T:3383865877;N:1221970	10	10	28	90	3779370721	3059502508	3396295214	3383865877	1221970	SRX21332627	SRS18578259	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.94896	0.14559	0.8002	0.51418	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25115	25115	SRR25605436	SRX21332626	SRS18578258	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep2	GSM7702833		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702833	GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq	GSM7702833 r1	GSM7702833	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py\|options: allowEarlyFileEnd	1907_GFP_pos_S2_L001_I1_001.fastq.gz 1907_GFP_pos_S2_L001_I2_001.fastq.gz 1907_GFP_pos_S2_L001_R1_001.fastq.gz 1907_GFP_pos_S2_L001_R2_001.fastq.gz	fastq fastq fastq fastq	22174775364.0	160686778.0	GSM7702833 r1	0:10 1:10 2:28 3:90	A:4207510387;C:3050740898;G:3362227609;T:3839333241;N:1997885	10	10	28	90	4207510387	3050740898	3362227609	3839333241	1997885	SRX21332626	SRS18578258	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.93287	0.11844	0.81087	0.55264	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25116	25116	SRR25605437	SRX21332626	SRS18578258	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep2	GSM7702833		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702833	GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq	GSM7702833 r1	GSM7702833	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py\|options: allowEarlyFileEnd	1907_GFP_pos_S2_L002_I1_001.fastq.gz 1907_GFP_pos_S2_L002_I2_001.fastq.gz 1907_GFP_pos_S2_L002_R1_001.fastq.gz 1907_GFP_pos_S2_L002_R2_001.fastq.gz	fastq fastq fastq fastq	22047847518.0	159767011.0	GSM7702833 r2	0:10 1:10 2:28 3:90	A:4174013532;C:3033959926;G:3357962867;T:3813042664;N:52001	10	10	28	90	4174013532	3033959926	3357962867	3813042664	52001	SRX21332626	SRS18578258	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.93359	0.11585	0.81087	0.55705	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25117	25117	SRR25822228	SRX21332626	SRS18578258	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep2	GSM7702833		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702833	GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq	GSM7702833 r1	GSM7702833	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_GFP_pos_S2_L001_I1_002.fastq.gz 1907_GFP_pos_S2_L001_I2_002.fastq.gz 1907_GFP_pos_S2_L001_R1_002.fastq.gz 1907_GFP_pos_S2_L001_R2_002.fastq.gz	fastq fastq fastq fastq	22010894568.0	159499236.0	GSM7702833 r3	0:10 1:10 2:28 3:90	A:4176090595;C:3026940106;G:3336506731;T:3814128890;N:1264918	10	10	28	90	4176090595	3026940106	3336506731	3814128890	1264918	SRX21332626	SRS18578258	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.93457	0.11733	0.81032	0.55203	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25118	25118	SRR25822229	SRX21332626	SRS18578258	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		gfp enriched rep2	GSM7702833		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	gfp enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702833	GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq	GSM7702833 r1	GSM7702833	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_GFP_pos_S2_L002_I1_002.fastq.gz 1907_GFP_pos_S2_L002_I2_002.fastq.gz 1907_GFP_pos_S2_L002_R1_002.fastq.gz 1907_GFP_pos_S2_L002_R2_002.fastq.gz	fastq fastq fastq fastq	22227220746.0	161066817.0	GSM7702833 r4	0:10 1:10 2:28 3:90	A:4209198596;C:3055507768;G:3382717459;T:3847294849;N:1294858	10	10	28	90	4209198596	3055507768	3382717459	3847294849	1294858	SRX21332626	SRS18578258	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.93428	0.11629	0.81014	0.56014	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25119	25119	SRR25605438	SRX21332625	SRS18578257	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep2	GSM7702832		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702832	GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq	GSM7702832 r1	GSM7702832	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_pos_pos_S1_L001_I1_001.fastq.gz 1907_pos_pos_S1_L001_I2_001.fastq.gz 1907_pos_pos_S1_L001_R1_001.fastq.gz 1907_pos_pos_S1_L001_R2_001.fastq.gz	fastq fastq fastq fastq	25141955454.0	182188083.0	GSM7702832 r1	0:10 1:10 2:28 3:90	A:4942067104;C:3446689328;G:3778138998;T:4227670796;N:2361244	10	10	28	90	4942067104	3446689328	3778138998	4227670796	2361244	SRX21332625	SRS18578257	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.90803	0.11273	0.81872	0.54345	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25120	25120	SRR25605439	SRX21332625	SRS18578257	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep2	GSM7702832		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702832	GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq	GSM7702832 r1	GSM7702832	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_pos_pos_S1_L002_R2_001.fastq.gz 1907_pos_pos_S1_L002_R1_001.fastq.gz 1907_pos_pos_S1_L002_I2_001.fastq.gz 1907_pos_pos_S1_L002_I1_001.fastq.gz	fastq fastq fastq fastq	25433730786.0	184302397.0	GSM7702832 r2	0:10 1:10 2:28 3:90	A:4980326106;C:3488986366;G:3842594871;T:4275245265;N:63122	10	10	28	90	4980326106	3488986366	3842594871	4275245265	63122	SRX21332625	SRS18578257	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.91035	0.11175	0.82016	0.52814	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25121	25121	SRR25822226	SRX21332625	SRS18578257	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep2	GSM7702832		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702832	GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq	GSM7702832 r1	GSM7702832	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_pos_pos_S1_L001_R2_002.fastq.gz 1907_pos_pos_S1_L001_R1_002.fastq.gz 1907_pos_pos_S1_L001_I2_002.fastq.gz 1907_pos_pos_S1_L001_I1_002.fastq.gz	fastq fastq fastq fastq	24834656088.0	179961276.0	GSM7702832 r3	0:10 1:10 2:28 3:90	A:4878672093;C:3403443000;G:3732878174;T:4180051888;N:1469685	10	10	28	90	4878672093	3403443000	3732878174	4180051888	1469685	SRX21332625	SRS18578257	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.90776	0.11325	0.81852	0.54011	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
25122	25122	SRR25822227	SRX21332625	SRS18578257	SRP454539	PRJNA1004255	Unique activities of two overlapping PAX6 retinal enhancers	GSE240575	Transcriptome Analysis	Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.		pubmed:37643867		mcherry enriched rep2	GSM7702832		source name:Eye\|tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf\|geo loc name:missing\|collection date:missing	mcherry enriched rep2	Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted…	Eye	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.	tissue:Eye\|genotype:NRE eGFP/HS5 mCherry\|developmental stage:48 hpf	GSM7702832	GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq	GSM7702832 r1	GSM7702832	1	NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	NextSeq 2000	SRP454539		loader:fastq load.py	1907_pos_pos_S1_L002_I1_002.fastq.gz 1907_pos_pos_S1_L002_I2_002.fastq.gz 1907_pos_pos_S1_L002_R1_002.fastq.gz 1907_pos_pos_S1_L002_R2_002.fastq.gz	fastq fastq fastq fastq	25609437216.0	185575632.0	GSM7702832 r4	0:10 1:10 2:28 3:90	A:5015380345;C:3509305882;G:3867598814;T:4308011415;N:1510424	10	10	28	90	5015380345	3509305882	3867598814	4308011415	1510424	SRX21332625	SRS18578257	SRA1702612	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh	1	0.90907	0.11192	0.81994	0.50902	90	B	usable mapping rate	illumina	nextseq_v2	3prime	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United Kingdom	2023-08-10	Hatching	Embryo	Eye	Sensory System
26558	26558	SRR26173859	SRX21885960	SRS18977085	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F17 R1	GSM7804200		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F17 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804200	GSM7804200: V2a sample2 354 F17 R1; Danio rerio; RNA Seq	GSM7804200 r1	GSM7804200	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F17_R1.fastq.gz	fastq	26979490.0	627430.0	GSM7804200 r1	0:43	A:7493119;C:5872644;G:6013311;T:7600416;N:0	43				7493119	5872644	6013311	7600416	0	SRX21885960	SRS18977085	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.82979	0.33034	0.94253	0.53995	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26559	26559	SRR26173860	SRX21885959	SRS18977083	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F16 R1	GSM7804199		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F16 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804199	GSM7804199: V2a sample2 354 F16 R1; Danio rerio; RNA Seq	GSM7804199 r1	GSM7804199	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F16_R1.fastq.gz	fastq	26312130.0	611910.0	GSM7804199 r1	0:43	A:7117249;C:5957278;G:6103577;T:7134026;N:0	43				7117249	5957278	6103577	7134026	0	SRX21885959	SRS18977083	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.87071	0.23579	0.90678	0.52226	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26560	26560	SRR26173861	SRX21885958	SRS18977084	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F15 R1	GSM7804198		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F15 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804198	GSM7804198: V2a sample2 354 F15 R1; Danio rerio; RNA Seq	GSM7804198 r1	GSM7804198	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F15_R1.fastq.gz	fastq	32073872.0	745904.0	GSM7804198 r1	0:43	A:8695962;C:7255833;G:7422516;T:8699561;N:0	43				8695962	7255833	7422516	8699561	0	SRX21885958	SRS18977084	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.86262	0.24391	0.90881	0.52511	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26561	26561	SRR26173862	SRX21885957	SRS18977081	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F14 R1	GSM7804197		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F14 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804197	GSM7804197: V2a sample2 354 F14 R1; Danio rerio; RNA Seq	GSM7804197 r1	GSM7804197	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F14_R1.fastq.gz	fastq	31158875.0	724625.0	GSM7804197 r1	0:43	A:8769852;C:6707813;G:6866255;T:8814955;N:0	43				8769852	6707813	6866255	8814955	0	SRX21885957	SRS18977081	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.8739	0.27254	0.9246	0.54185	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26562	26562	SRR26173863	SRX21885956	SRS18977082	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F13 R1	GSM7804196		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F13 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804196	GSM7804196: V2a sample2 354 F13 R1; Danio rerio; RNA Seq	GSM7804196 r1	GSM7804196	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F13_R1.fastq.gz	fastq	41289804.0	960228.0	GSM7804196 r1	0:43	A:11114918;C:9358398;G:9568761;T:11247727;N:0	43				11114918	9358398	9568761	11247727	0	SRX21885956	SRS18977082	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.82861	0.29409	0.93235	0.49485	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26563	26563	SRR26173864	SRX21885955	SRS18977079	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F12 R1	GSM7804195		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F12 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804195	GSM7804195: V2a sample2 354 F12 R1; Danio rerio; RNA Seq	GSM7804195 r1	GSM7804195	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F12_R1.fastq.gz	fastq	30569259.0	710913.0	GSM7804195 r1	0:43	A:8286806;C:6854804;G:7028453;T:8399196;N:0	43				8286806	6854804	7028453	8399196	0	SRX21885955	SRS18977079	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.80426	0.29136	0.94123	0.54305	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26564	26564	SRR26173865	SRX21885954	SRS18977080	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F11 R1	GSM7804194		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F11 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804194	GSM7804194: V2a sample2 354 F11 R1; Danio rerio; RNA Seq	GSM7804194 r1	GSM7804194	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F11_R1.fastq.gz	fastq	29626097.0	688979.0	GSM7804194 r1	0:43	A:8063765;C:6645347;G:6803778;T:8113207;N:0	43				8063765	6645347	6803778	8113207	0	SRX21885954	SRS18977080	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.84972	0.26423	0.9234	0.54087	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26565	26565	SRR26173866	SRX21885953	SRS18977078	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 F10 R1	GSM7804193		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 F10 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804193	GSM7804193: V2a sample2 354 F10 R1; Danio rerio; RNA Seq	GSM7804193 r1	GSM7804193	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_F10_R1.fastq.gz	fastq	24594409.0	571963.0	GSM7804193 r1	0:43	A:6605642;C:5535165;G:5682586;T:6771016;N:0	43				6605642	5535165	5682586	6771016	0	SRX21885953	SRS18977078	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.77683	0.28872	0.94633	0.52513	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26566	26566	SRR26173867	SRX21885952	SRS18977077	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E9 R1	GSM7804168		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E9 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804168	GSM7804168: V2a sample2 354 E9 R1; Danio rerio; RNA Seq	GSM7804168 r1	GSM7804168	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E9_R1.fastq.gz	fastq	33259941.0	773487.0	GSM7804168 r1	0:43	A:9004615;C:7506729;G:7674909;T:9073688;N:0	43				9004615	7506729	7674909	9073688	0	SRX21885952	SRS18977077	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.85528	0.32094	0.89217	0.50224	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26567	26567	SRR26173868	SRX21885951	SRS18977076	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E8 R1	GSM7804167		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E8 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804167	GSM7804167: V2a sample2 354 E8 R1; Danio rerio; RNA Seq	GSM7804167 r1	GSM7804167	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E8_R1.fastq.gz	fastq	45348316.0	1054612.0	GSM7804167 r1	0:43	A:12143243;C:10459395;G:10612458;T:12133220;N:0	43				12143243	10459395	10612458	12133220	0	SRX21885951	SRS18977076	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.8783	0.19318	0.88572	0.49101	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26568	26568	SRR26173869	SRX21885950	SRS18977075	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E7 R1	GSM7804166		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E7 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804166	GSM7804166: V2a sample2 354 E7 R1; Danio rerio; RNA Seq	GSM7804166 r1	GSM7804166	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E7_R1.fastq.gz	fastq	30522002.0	709814.0	GSM7804166 r1	0:43	A:8167984;C:7033955;G:7127866;T:8192197;N:0	43				8167984	7033955	7127866	8192197	0	SRX21885950	SRS18977075	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.87804	0.20762	0.88418	0.51381	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26569	26569	SRR26173870	SRX21885949	SRS18977074	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E6 R1	GSM7804165		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E6 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804165	GSM7804165: V2a sample2 354 E6 R1; Danio rerio; RNA Seq	GSM7804165 r1	GSM7804165	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E6_R1.fastq.gz	fastq	39095858.0	909206.0	GSM7804165 r1	0:43	A:10597068;C:8804712;G:8974908;T:10719170;N:0	43				10597068	8804712	8974908	10719170	0	SRX21885949	SRS18977074	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.8093	0.38297	0.91545	0.56195	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26570	26570	SRR26173871	SRX21885948	SRS18977073	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E5 R1	GSM7804164		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E5 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804164	GSM7804164: V2a sample2 354 E5 R1; Danio rerio; RNA Seq	GSM7804164 r1	GSM7804164	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E5_R1.fastq.gz	fastq	33632880.0	782160.0	GSM7804164 r1	0:43	A:9099447;C:7602241;G:7747634;T:9183558;N:0	43				9099447	7602241	7747634	9183558	0	SRX21885948	SRS18977073	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.83226	0.28405	0.90715	0.52148	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26571	26571	SRR26173872	SRX21885947	SRS18977072	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E4 R1	GSM7804163		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E4 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804163	GSM7804163: V2a sample2 354 E4 R1; Danio rerio; RNA Seq	GSM7804163 r1	GSM7804163	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E4_R1.fastq.gz	fastq	10823057.0	251699.0	GSM7804163 r1	0:43	A:3014035;C:2419189;G:2487845;T:2901988;N:0	43				3014035	2419189	2487845	2901988	0	SRX21885947	SRS18977072	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.85783	0.26946	0.91033	0.52459	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26572	26572	SRR26173873	SRX21885946	SRS18977068	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E3 R1	GSM7804162		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E3 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804162	GSM7804162: V2a sample2 354 E3 R1; Danio rerio; RNA Seq	GSM7804162 r1	GSM7804162	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E3_R1.fastq.gz	fastq	49775381.0	1157567.0	GSM7804162 r1	0:43	A:13325978;C:11313973;G:11568082;T:13567348;N:0	43				13325978	11313973	11568082	13567348	0	SRX21885946	SRS18977068	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.80873	0.29504	0.93791	0.52789	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26573	26573	SRR26173874	SRX21885945	SRS18977070	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 E2 R1	GSM7804161		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 E2 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804161	GSM7804161: V2a sample2 354 E2 R1; Danio rerio; RNA Seq	GSM7804161 r1	GSM7804161	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_E2_R1.fastq.gz	fastq	44072334.0	1024938.0	GSM7804161 r1	0:43	A:11780930;C:10068667;G:10185567;T:12037170;N:0	43				11780930	10068667	10185567	12037170	0	SRX21885945	SRS18977070	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.78009	0.2621	0.943	0.54719	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26574	26574	SRR26173875	SRX21885944	SRS18977071	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 D1 R1	GSM7804136		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 D1 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804136	GSM7804136: V2a sample2 354 D1 R1; Danio rerio; RNA Seq	GSM7804136 r1	GSM7804136	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_D1_R1.fastq.gz	fastq	49923.0	1161.0	GSM7804136 r1	0:43	A:12799;C:11604;G:11134;T:14386;N:0	43				12799	11604	11134	14386	0	SRX21885944	SRS18977071	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.5773	0.18478	0.99472	0.51097	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26575	26575	SRR26173876	SRX21885943	SRS18977069	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C24 R1	GSM7804135		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C24 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804135	GSM7804135: V2a sample2 354 C24 R1; Danio rerio; RNA Seq	GSM7804135 r1	GSM7804135	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C24_R1.fastq.gz	fastq	30735540.0	714780.0	GSM7804135 r1	0:43	A:7999390;C:7036767;G:7190113;T:8509270;N:0	43				7999390	7036767	7190113	8509270	0	SRX21885943	SRS18977069	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.48457	0.16974	0.99328	0.82979	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26576	26576	SRR26173877	SRX21885942	SRS18977067	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C23 R1	GSM7804134		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C23 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804134	GSM7804134: V2a sample2 354 C23 R1; Danio rerio; RNA Seq	GSM7804134 r1	GSM7804134	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C23_R1.fastq.gz	fastq	24386031.0	567117.0	GSM7804134 r1	0:43	A:6620599;C:5230020;G:5371347;T:7164065;N:0	43				6620599	5230020	5371347	7164065	0	SRX21885942	SRS18977067	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.55183	0.13531	0.99105	0.54904	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26577	26577	SRR26173878	SRX21885941	SRS18977064	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C22 R1	GSM7804133		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C22 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804133	GSM7804133: V2a sample2 354 C22 R1; Danio rerio; RNA Seq	GSM7804133 r1	GSM7804133	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C22_R1.fastq.gz	fastq	35984550.0	836850.0	GSM7804133 r1	0:43	A:9917644;C:7959027;G:8126203;T:9981676;N:0	43				9917644	7959027	8126203	9981676	0	SRX21885941	SRS18977064	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.85473	0.2804	0.91165	0.52909	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26578	26578	SRR26173879	SRX21885940	SRS18977065	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C21 R1	GSM7804132		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C21 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804132	GSM7804132: V2a sample2 354 C21 R1; Danio rerio; RNA Seq	GSM7804132 r1	GSM7804132	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C21_R1.fastq.gz	fastq	32265738.0	750366.0	GSM7804132 r1	0:43	A:8963112;C:7046873;G:7192067;T:9063686;N:0	43				8963112	7046873	7192067	9063686	0	SRX21885940	SRS18977065	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.8426	0.36876	0.91179	0.58759	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26579	26579	SRR26173880	SRX21885939	SRS18977066	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C20 R1	GSM7804131		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C20 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804131	GSM7804131: V2a sample2 354 C20 R1; Danio rerio; RNA Seq	GSM7804131 r1	GSM7804131	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C20_R1.fastq.gz	fastq	33471157.0	778399.0	GSM7804131 r1	0:43	A:9073629;C:7510863;G:7675381;T:9211284;N:0	43				9073629	7510863	7675381	9211284	0	SRX21885939	SRS18977066	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.82141	0.28926	0.93655	0.49873	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26580	26580	SRR26173881	SRX21885938	SRS18977062	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C19 R1	GSM7804130		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C19 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804130	GSM7804130: V2a sample2 354 C19 R1; Danio rerio; RNA Seq	GSM7804130 r1	GSM7804130	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C19_R1.fastq.gz	fastq	30131304.0	700728.0	GSM7804130 r1	0:43	A:8036932;C:6893678;G:7034855;T:8165839;N:0	43				8036932	6893678	7034855	8165839	0	SRX21885938	SRS18977062	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.80896	0.25245	0.94067	0.54072	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26581	26581	SRR26173882	SRX21885937	SRS18977061	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 C18 R1	GSM7804129		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 C18 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804129	GSM7804129: V2a sample2 354 C18 R1; Danio rerio; RNA Seq	GSM7804129 r1	GSM7804129	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_C18_R1.fastq.gz	fastq	31050429.0	722103.0	GSM7804129 r1	0:43	A:8363024;C:7088111;G:7227301;T:8371993;N:0	43				8363024	7088111	7227301	8371993	0	SRX21885937	SRS18977061	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.87399	0.20843	0.89881	0.49682	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26582	26582	SRR26173883	SRX21885936	SRS18977063	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 B17 R1	GSM7804104		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 B17 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804104	GSM7804104: V2a sample2 354 B17 R1; Danio rerio; RNA Seq	GSM7804104 r1	GSM7804104	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_B17_R1.fastq.gz	fastq	47093729.0	1095203.0	GSM7804104 r1	0:43	A:13057598;C:10185883;G:10457076;T:13393172;N:0	43				13057598	10185883	10457076	13393172	0	SRX21885936	SRS18977063	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.80343	0.36098	0.95059	0.52562	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26583	26583	SRR26173884	SRX21885935	SRS18977060	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 B16 R1	GSM7804103		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 B16 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804103	GSM7804103: V2a sample2 354 B16 R1; Danio rerio; RNA Seq	GSM7804103 r1	GSM7804103	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_B16_R1.fastq.gz	fastq	34160791.0	794437.0	GSM7804103 r1	0:43	A:9178469;C:7729053;G:7901331;T:9351938;N:0	43				9178469	7729053	7901331	9351938	0	SRX21885935	SRS18977060	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.79028	0.25695	0.94627	0.51668	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26584	26584	SRR26173885	SRX21885934	SRS18977059	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 B15 R1	GSM7804102		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 B15 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804102	GSM7804102: V2a sample2 354 B15 R1; Danio rerio; RNA Seq	GSM7804102 r1	GSM7804102	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_B15_R1.fastq.gz	fastq	29152495.0	677965.0	GSM7804102 r1	0:43	A:7966546;C:6516464;G:6675123;T:7994362;N:0	43				7966546	6516464	6675123	7994362	0	SRX21885934	SRS18977059	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.83832	0.25969	0.92673	0.51661	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26585	26585	SRR26173886	SRX21885933	SRS18977057	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 B14 R1	GSM7804101		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 B14 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804101	GSM7804101: V2a sample2 354 B14 R1; Danio rerio; RNA Seq	GSM7804101 r1	GSM7804101	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_B14_R1.fastq.gz	fastq	23811379.0	553753.0	GSM7804101 r1	0:43	A:6532691;C:5215122;G:5353003;T:6710563;N:0	43				6532691	5215122	5353003	6710563	0	SRX21885933	SRS18977057	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.74067	0.3282	0.95272	0.49582	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System
26586	26586	SRR26173887	SRX21885932	SRS18977058	SRP463130	PRJNA1020854	Molecular blueprints for spinal circuit modules controlling locomotor speed	GSE243993	Transcriptome Analysis	The flexibility of motor actions is ingrained in the diversity of neurons and how they are organized into functional circuit modules yet our knowledge of the molecular underpinning of motor circuit modularity remains limited. Locomotion is a motor behavior characterized by sudden changes in speed and strength enabled by the coordinated recruitment of different motoneuron subtypes. Here we use adult zebrafish to link the molecular diversity of motoneurons and the rhythm generating V2a interneurons with their modular circuit organization that is responsible for changes in locomotor speed. We show that the molecular diversity of motoneurons and V2a interneurons reflects their functional segregation into slow intermediate or fast subtypes. Furthermore we reveal shared molecular signatures between V2a interneurons and motoneurons of the three speed circuit modules. Overall by characterizing how the molecular diversity of motoneurons and V2a interneurons relates to their function connectivity and behavior our study provides important insights not only into the molecular mechanisms for neuronal and circuit diversity for locomotor flexibility but also for charting circuits for motor actions in general. Overall design: To determine whether the functional subtypes of motoneurons and V2a Chx10+ interneurons are molecularly distinct we performed single cell RNA sequencing respectively on adult islet1a:GFP and chx10:GFP transgenic zebrafish using SmartSeq2.		pubmed:37919423		V2a sample2 354 B13 R1	GSM7804100		source name:Spinal cord\|tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons\|geo loc name:missing\|collection date:missing	V2a sample2 354 B13 R1	The reads from each sequenced cell were mapped to the zebrafish reference genome “Danio rerio Ensembl GRCz11” using STAR version 2.5.3a. The resulting bam files were filtered to keep only uniquely mapped reads. Most of the following analysis was performed in R version 4.0.5 R core team 2022 using the Seurat package version 4.0.2. Assembly: GRCz11 Supplementary files format and content: .csv files with gene count matrixes; .txt and .csv metadata files and .rds files containing R objects from Seurat analysis	Spinal cord	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2	tissue:Spinal cord\|cell line:Chx10:GFP\|cell type:V2a interneurons	GSM7804100	GSM7804100: V2a sample2 354 B13 R1; Danio rerio; RNA Seq	GSM7804100 r1	GSM7804100	1	Adult animals 7 wpf of either sex were deeply anesthetized in a slush of frozen extracellular solution containing in mM: 134 NaCl 2.9 KCl 2.1 CaCl2 1.2 MgCl2 10 HEPES and 10 glucose with pH of 7.8 adjusted with NaOH and osmolarity of 290 mOsm. The spinal cord was quickly dissected in the slush of frozen extracellular solution and collected. Two samples were prepared from the Tgislet1a:GFP line and two samples were prepared from the Tgchx10:GFP line. For each sample 6 to 10 intact isolated spinal cords were incubated in 1 ml of DMEM F12 medium Thermo Fisher #11039021 osmolarity adjusted to 280 280 mOsm containing papain 10 U/ml Worthington biochem #LK003178 on a heated shaker at 37°C for 15 min. DMEM/F 12 1 ml 280 290 mOsm was added to stop the enzymatic reaction. The sample was centrifuged at 300 g at 4°C for 5 min and then re suspended in 0.5 ml of DMEM/F 12 280 290 mOsm post removal of the supernatant. Following mechanical trituration using fire polished Pasteur pipettes the cell suspension was filtered through a cell 16 strainer 40 μm. The sample was kept at room temperature for 20 min post the addition of 0 1 ml of the nuclear DNA stain DRAQ5 Thermo Fisher #65 0880 92. Using fluorescence activated cell sorting FACs cells positive for GFP and DRAQ5 in each sample were sorted into a 384 wells plate containing a mild hypotonic lysis buffer 0.2% Triton X 100 2 U/ml RNase inhibitor and immediately snap frozen on ice then stored at 80°C. Smart Seq2		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	SINGLE	ILLUMINA	Illumina HiSeq 2000	SRP463130			SS2_18_354_B13_R1.fastq.gz	fastq	18141356.0	421892.0	GSM7804100 r1	0:43	A:4926550;C:4092334;G:4186281;T:4936191;N:0	43				4926550	4092334	4186281	4936191	0	SRX21885932	SRS18977058	SRA1719948	Neuroscience, Karolinaska Institutet	Neuroscience, Karolinaska Institutet	1	0.87463	0.22642	0.89826	0.50541	43	B	usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_plate	smartseq	Sweden	2023-09-25	Juvenile	Juvenile	Spinal Cord	Nervous System

Advanced export

JSON shape: default, array, newline-delimited

CREATE TABLE run_metadata("run.accession" VARCHAR, "experiment.accession" VARCHAR, "sample.accession" VARCHAR, "study.accession" VARCHAR, bioproject VARCHAR, "study.title" VARCHAR, "study.alias" VARCHAR, "study.type" VARCHAR, "study.abstract" VARCHAR, "study.attributes" VARCHAR, "study.PMIDs" VARCHAR, "sample.description" VARCHAR, "sample.title" VARCHAR, "sample.alias" VARCHAR, "sample.centername" VARCHAR, "sample.attributes" VARCHAR, "GEOsample.title" VARCHAR, "GEOsample.dataprocessing" VARCHAR, "GEOsample.source" VARCHAR, "GEOsample.treatmentprotocol" VARCHAR, "GEOsample.extractprotocol" VARCHAR, "GEOsample.growthprotocol" VARCHAR, "GEOsample.characteristics" VARCHAR, "GEOsample.accession" VARCHAR, "experiment.title" VARCHAR, "experiment.alias" VARCHAR, "experiment.library_name" VARCHAR, "experiment.design_description" VARCHAR, "experiment.library_construction_protocol" VARCHAR, "experiment.attributes" VARCHAR, "experiment.library_strategy" VARCHAR, "experiment.library_source" VARCHAR, "experiment.library_selection" VARCHAR, "experiment.library_layout" VARCHAR, "experiment.platform" VARCHAR, "experiment.instrument_model" VARCHAR, "experiment.spot_descriptor" VARCHAR, "experiment.study_ref" VARCHAR, "run.title" VARCHAR, "run.attributes" VARCHAR, "run.filename" VARCHAR, "run.semantic_name" VARCHAR, "run.total_bases" DOUBLE, "run.total_spots" DOUBLE, "run.alias" VARCHAR, "run.read_lengths" VARCHAR, "run.base_counts" VARCHAR, "run.r1_length" BIGINT, "run.r2_length" BIGINT, "run.r3_length" BIGINT, "run.r4_length" BIGINT, "run.Acount" BIGINT, "run.Ccount" BIGINT, "run.Gcount" BIGINT, "run.Tcount" BIGINT, "run.Ncount" BIGINT, "run.experiment" VARCHAR, "run.pool_member" VARCHAR, "submission.accession" VARCHAR, "submission.srasource" VARCHAR, "submission.bioprojectsource" VARCHAR, "seqdetective.n_mates" BIGINT, "seqdetective.mapping_rate.mate1" DOUBLE, "seqdetective.mapping_rate.mate2" DOUBLE, "seqdetective.nofeature_rate.mate1" DOUBLE, "seqdetective.nofeature_rate.mate2" DOUBLE, "seqdetective.sparsity.mate1" DOUBLE, "seqdetective.sparsity.mate2" DOUBLE, "seqdetective.pos_strand_rate.mate1" DOUBLE, "seqdetective.pos_strand_rate.mate2" DOUBLE, "seqdetective.readlen.mate1" BIGINT, "seqdetective.readlen.mate2" BIGINT, "seqdetective.judgement.mate1" VARCHAR, "seqdetective.judgement.mate2" VARCHAR, "seqdetective.judgement.reason" VARCHAR, platform_family VARCHAR, instrument_generation VARCHAR, read_bias VARCHAR, selection_class VARCHAR, prep_kit VARCHAR, sc_or_bulk VARCHAR, tech_class VARCHAR, technology VARCHAR, tech_variant VARCHAR, "submission.bioprojectsource.country" VARCHAR, earliest_date DATE, devstage_curation VARCHAR, devstage_curation_coarse VARCHAR, tissue_curation VARCHAR, tissue_curation_coarse VARCHAR);;
CREATE INDEX idx_run_bioproject ON run_metadata(bioproject);;
CREATE INDEX idx_run_run_accession ON run_metadata("run.accession");;