metadata: run_metadata: 63 rows where experiment.platform = "ILLUMINA", technology = "10x" and tissue

63 rows where experiment.platform = "ILLUMINA", technology = "10x" and tissue_curation = "Spinal Cord"

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.treatmentprotocol	GEOsample.extractprotocol	GEOsample.growthprotocol	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.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.mapping_rate.mate2	seqdetective.nofeature_rate.mate1	seqdetective.nofeature_rate.mate2	seqdetective.sparsity.mate1	seqdetective.sparsity.mate2	seqdetective.pos_strand_rate.mate1	seqdetective.pos_strand_rate.mate2	seqdetective.readlen.mate1	seqdetective.readlen.mate2	seqdetective.judgement.mate1	seqdetective.judgement.mate2	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
10176	10176	ERR5858457	ERX5504346	ERS6343450	ERP128749	PRJEB44676	scRNAseq of her4.3+ cells from lesioned and unlesioned zebrafish larvae spinal cord	E-MTAB-10390	Transcriptome Analysis	To analyse lesion induced gene regulation in progenitor cells at single cell resolution we performed single cell RNAseq on FACS isolated her4.3:GFP progenitor 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 05 24\|ENA LAST UPDATE:2021 05 24		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	SAMEA8658904	University Of Edinburgh	ENA first public:2021 05 24\|ENA last update:2021 05 24\|External Id:SAMEA8658904\|INSDC center alias:UOE\|INSDC center name:University Of Edinburgh\|INSDC first public:2021 05 24T00:14:32Z\|INSDC last update:2021 05 24T00:14:32Z\|INSDC status:public\|Submitter Id:E MTAB 10390:Naive\|age:4\|broker name:ArrayExpress\|cell type:ependymo radial glial cell\|common name:zebrafish\|developmental stage:larval day 4\|immunophenotype:Her4.3+ positive\|organism part:spinal cord\|sample name:E MTAB 10390:Naive\|sex:mixed\|strain:WIK									Illumina NovaSeq 6000 paired end sequencing; scRNAseq of her4.3+ cells from lesi1d and unlesi1d zebrafish larvae spinal cord	E MTAB 10390:Naive p	Naive p	scRNAseq of her4.3+ 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	PAIRED	ILLUMINA	Illumina NovaSeq 6000	ERP128749	Illumina NovaSeq 6000 paired end sequencing; scRNAseq of her4.3+ cells from lesioned and unlesioned zebrafish larvae spinal cord	ENA FIRST PUBLIC:2021 05 24\|ENA LAST UPDATE:2021 05 24	Naive.bam	bam	44972162730.0	499690697.0	E MTAB 10390:Naive	0:90	A:13658826375;C:8733304343;G:9348137569;T:13228064343;N:3830100	90			13658826375	8733304343	9348137569	13228064343	3830100	ERX5504346	ERS6343450	ERA4142789	University Of Edinburgh\|European Nucleotide Archive	University Of Edinburgh\|European Nucleotide Archive	1	0.89428		0.32693		0.75276		0.5314		90		B		usable mapping rate	illumina	novaseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	United Kingdom	2021-05-24	Larval	Larval	Spinal Cord	Nervous System
10177	10177	ERR5858456	ERX5504345	ERS6343449	ERP128749	PRJEB44676	scRNAseq of her4.3+ cells from lesioned and unlesioned zebrafish larvae spinal cord	E-MTAB-10390	Transcriptome Analysis	To analyse lesion induced gene regulation in progenitor cells at single cell resolution we performed single cell RNAseq on FACS isolated her4.3:GFP progenitor 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 05 24\|ENA LAST UPDATE:2021 05 24		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	SAMEA8658903	University Of Edinburgh	ENA first public:2021 05 24\|ENA last update:2021 05 24\|External Id:SAMEA8658903\|INSDC center alias:UOE\|INSDC center name:University Of Edinburgh\|INSDC first public:2021 05 24T00:14:32Z\|INSDC last update:2021 05 24T00:14:32Z\|INSDC status:public\|Submitter Id:E MTAB 10390:Lesi1d\|age:4\|broker name:ArrayExpress\|cell type:ependymo radial glial cell\|common name:zebrafish\|developmental stage:larval day 4\|immunophenotype:Her4.3+ positive\|injury:spinal injury lesion\|organism part:spinal cord\|sample name:E MTAB 10390:Lesi1d\|sex:mixed\|strain:WIK									Illumina NovaSeq 6000 paired end sequencing; scRNAseq of her4.3+ cells from lesi1d and unlesi1d zebrafish larvae spinal cord	E MTAB 10390:Lesioned p	Lesioned p	scRNAseq of her4.3+ 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 injury lesion	RNA-Seq	TRANSCRIPTOMIC	Oligo-dT	PAIRED	ILLUMINA	Illumina NovaSeq 6000	ERP128749	Illumina NovaSeq 6000 paired end sequencing; scRNAseq of her4.3+ cells from lesioned and unlesioned zebrafish larvae spinal cord	ENA FIRST PUBLIC:2021 05 24\|ENA LAST UPDATE:2021 05 24	Lesioned.bam	bam	49902588630.0	554473207.0	E MTAB 10390:Lesioned	0:90	A:14713351178;C:10191139050;G:10897887675;T:14095967201;N:4243526	90			14713351178	10191139050	10897887675	14095967201	4243526	ERX5504345	ERS6343449	ERA4142789	University Of Edinburgh\|European Nucleotide Archive	University Of Edinburgh\|European Nucleotide Archive	1	0.91197		0.29137		0.7568		0.56523		90		B		usable mapping rate	illumina	novaseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x	United Kingdom	2021-05-24	Larval	Larval	Spinal Cord	Nervous System
24967	24967	SRR25557924	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5_S3_L001_I1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L001_R1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L001_R2_001.fastq.gz	fastq fastq fastq	783833459.0	6171917.0	GSM7688796 r1	0:8 1:28 2:91	A:165863131;C:117191802;G:128146802;T:150173329;N:269383	8	28	91	165863131	117191802	128146802	150173329	269383	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89127		0.221		0.78317		0.52195		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24968	24968	SRR25557925	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-3_S2_L002_I1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L002_R1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L002_R2_001.fastq.gz	fastq fastq fastq	7208342327.0	56758601.0	GSM7688796 r10	0:8 1:28 2:91	A:1518008610;C:1083228714;G:1190089577;T:1369772782;N:3933008	8	28	91	1518008610	1083228714	1190089577	1369772782	3933008	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.88938		0.21991		0.79088		0.53044		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24969	24969	SRR25557926	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-3_S2_L003_I1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L003_R1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L003_R2_001.fastq.gz	fastq fastq fastq	7337435414.0	57775082.0	GSM7688796 r11	0:8 1:28 2:91	A:1545051667;C:1103394058;G:1211577972;T:1395133564;N:2375201	8	28	91	1545051667	1103394058	1211577972	1395133564	2375201	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.88966		0.2211		0.78961		0.53156		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24970	24970	SRR25557927	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-3_S2_L004_I1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L004_R1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L004_R2_001.fastq.gz	fastq fastq fastq	7218240453.0	56836539.0	GSM7688796 r12	0:8 1:28 2:91	A:1521442237;C:1084242714;G:1191469584;T:1372277539;N:2692975	8	28	91	1521442237	1084242714	1191469584	1372277539	2692975	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.8888		0.2174		0.79172		0.53311		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24971	24971	SRR25557928	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5_S3_L002_I1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L002_R1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L002_R2_001.fastq.gz	fastq fastq fastq	774302109.0	6096867.0	GSM7688796 r2	0:8 1:28 2:91	A:163989105;C:115793536;G:126358958;T:148430473;N:242825	8	28	91	163989105	115793536	126358958	148430473	242825	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89371		0.2215		0.78253		0.53282		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24972	24972	SRR25557929	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5_S3_L003_I1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L003_R1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L003_R2_001.fastq.gz	fastq fastq fastq	790430093.0	6223859.0	GSM7688796 r3	0:8 1:28 2:91	A:167465790;C:118269595;G:129024496;T:151447225;N:164063	8	28	91	167465790	118269595	129024496	151447225	164063	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89284		0.21955		0.78356		0.53369		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24973	24973	SRR25557930	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5_S3_L004_I1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L004_R1_001.fastq.gz MUTSU2_AllGFP-S5_S3_L004_R2_001.fastq.gz	fastq fastq fastq	778936974.0	6133362.0	GSM7688796 r4	0:8 1:28 2:91	A:165013558;C:116498474;G:127146290;T:149314806;N:162814	8	28	91	165013558	116498474	127146290	149314806	162814	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89195		0.2209		0.78196		0.53121		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24974	24974	SRR25557931	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-2_S2_L001_I1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L001_R1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L001_R2_001.fastq.gz	fastq fastq fastq	4174244890.0	32868070.0	GSM7688796 r5	0:8 1:28 2:91	A:882060228;C:627098568;G:684890393;T:795503862;N:1441319	8	28	91	882060228	627098568	684890393	795503862	1441319	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89137		0.22041		0.77926		0.52991		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24975	24975	SRR25557932	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-2_S2_L002_I1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L002_R1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L002_R2_001.fastq.gz	fastq fastq fastq	4150210902.0	32678826.0	GSM7688796 r6	0:8 1:28 2:91	A:878218072;C:623662333;G:679977758;T:790589138;N:1325865	8	28	91	878218072	623662333	679977758	790589138	1325865	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.89264		0.22159		0.78121		0.52989		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24976	24976	SRR25557933	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-2_S2_L003_I1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L003_R1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L003_R2_001.fastq.gz	fastq fastq fastq	4198043547.0	33055461.0	GSM7688796 r7	0:8 1:28 2:91	A:888287475;C:630914324;G:688006958;T:799794077;N:1044117	8	28	91	888287475	630914324	688006958	799794077	1044117	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.8912		0.21889		0.7822		0.52774		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24977	24977	SRR25557934	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-2_S2_L004_I1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L004_R1_001.fastq.gz MUTSU2_AllGFP-S5-2_S2_L004_R2_001.fastq.gz	fastq fastq fastq	4154784553.0	32714839.0	GSM7688796 r8	0:8 1:28 2:91	A:880520885;C:624179045;G:680076172;T:791297938;N:976309	8	28	91	880520885	624179045	680076172	791297938	976309	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.8927		0.22063		0.78216		0.5346		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24978	24978	SRR25557935	SRX21286763	SRS18536876	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 AllGFP S5	GSM7688796		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 AllGFP S5	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688796	GSM7688796: MUTSU2 AllGFP S5; Danio rerio; RNA Seq	GSM7688796 r1	GSM7688796	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_AllGFP-S5-3_S2_L001_I1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L001_R1_001.fastq.gz MUTSU2_AllGFP-S5-3_S2_L001_R2_001.fastq.gz	fastq fastq fastq	7231973725.0	56944675.0	GSM7688796 r9	0:8 1:28 2:91	A:1522035629;C:1085783001;G:1195535350;T:1374885853;N:3725592	8	28	91	1522035629	1085783001	1195535350	1374885853	3725592	SRX21286763	SRS18536876	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.88833		0.22076		0.79056		0.52744		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24979	24979	SRR25557936	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8_S2_L001_I1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L001_R1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L001_R2_001.fastq.gz	fastq fastq fastq	699161670.0	5505210.0	GSM7688795 r1	0:8 1:28 2:91	A:149927051;C:100829317;G:116075103;T:133908276;N:234363	8	28	91	149927051	100829317	116075103	133908276	234363	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85081		0.21257		0.80426		0.52779		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24980	24980	SRR25557937	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-3_S1_L002_I1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L002_R1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L002_R2_001.fastq.gz	fastq fastq fastq	8174836330.0	64368790.0	GSM7688795 r10	0:8 1:28 2:91	A:1748532358;C:1187854561;G:1356279755;T:1560491610;N:4401606	8	28	91	1748532358	1187854561	1356279755	1560491610	4401606	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85069		0.20989		0.81049		0.52069		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24981	24981	SRR25557938	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-3_S1_L003_I1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L003_R1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L003_R2_001.fastq.gz	fastq fastq fastq	8307872640.0	65416320.0	GSM7688795 r11	0:8 1:28 2:91	A:1778112295;C:1207569588;G:1377139686;T:1587359299;N:2704252	8	28	91	1778112295	1207569588	1377139686	1587359299	2704252	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85151		0.21054		0.80813		0.52675		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24982	24982	SRR25557939	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-3_S1_L004_R2_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L004_R1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L004_I1_001.fastq.gz	fastq fastq fastq	8197523483.0	64547429.0	GSM7688795 r12	0:8 1:28 2:91	A:1755321680;C:1190415496;G:1359502433;T:1565500310;N:3076120	8	28	91	1755321680	1190415496	1359502433	1565500310	3076120	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.84956		0.20968		0.81113		0.52382		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24983	24983	SRR25557940	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8_S2_L002_R2_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L002_R1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L002_I1_001.fastq.gz	fastq fastq fastq	690198899.0	5434637.0	GSM7688795 r2	0:8 1:28 2:91	A:147896108;C:99606794;G:114681063;T:132140792;N:227210	8	28	91	147896108	99606794	114681063	132140792	227210	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85228		0.21261		0.803		0.52382		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24984	24984	SRR25557941	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8_S2_L003_I1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L003_R1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L003_R2_001.fastq.gz	fastq fastq fastq	702239515.0	5529445.0	GSM7688795 r3	0:8 1:28 2:91	A:150477574;C:101266550;G:116878466;T:134415299;N:141606	8	28	91	150477574	101266550	116878466	134415299	141606	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85364		0.21344		0.80346		0.52158		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24985	24985	SRR25557942	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8_S2_L004_I1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L004_R1_001.fastq.gz MUTSU2_OtherGFP-G8_S2_L004_R2_001.fastq.gz	fastq fastq fastq	694537346.0	5468798.0	GSM7688795 r4	0:8 1:28 2:91	A:148876263;C:100202441;G:115455212;T:132982029;N:144673	8	28	91	148876263	100202441	115455212	132982029	144673	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85238		0.2138		0.80379		0.52691		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24986	24986	SRR25557943	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-2_S1_L001_R2_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L001_R1_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L001_I1_001.fastq.gz	fastq fastq fastq	4324997954.0	34055102.0	GSM7688795 r5	0:8 1:28 2:91	A:924471384;C:626141120;G:722856562;T:824050239;N:1494977	8	28	91	924471384	626141120	722856562	824050239	1494977	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85083		0.21122		0.80472		0.524		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24987	24987	SRR25557944	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-2_S1_L002_R2_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L002_R1_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L002_I1_001.fastq.gz	fastq fastq fastq	4306232942.0	33907346.0	GSM7688795 r6	0:8 1:28 2:91	A:920615353;C:623090513;G:721201055;T:819291149;N:1370416	8	28	91	920615353	623090513	721201055	819291149	1370416	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85335		0.21315		0.80206		0.51508		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24988	24988	SRR25557945	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-2_S1_L003_R2_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L003_R1_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L003_I1_001.fastq.gz	fastq fastq fastq	4348436312.0	34239656.0	GSM7688795 r7	0:8 1:28 2:91	A:930204208;C:629005094;G:727756813;T:827747478;N:1095103	8	28	91	930204208	629005094	727756813	827747478	1095103	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85369		0.21216		0.80503		0.52219		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24989	24989	SRR25557946	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-2_S1_L004_R2_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L004_R1_001.fastq.gz MUTSU2_OtherGFP-G8-2_S1_L004_I1_001.fastq.gz	fastq fastq fastq	4313915172.0	33967836.0	GSM7688795 r8	0:8 1:28 2:91	A:922742754;C:623127831;G:724345885;T:819819593;N:1037013	8	28	91	922742754	623127831	724345885	819819593	1037013	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.85456		0.21236		0.80223		0.5216		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
24990	24990	SRR25557947	SRX21286762	SRS18536875	SRP453891	PRJNA1003032	Molecular Analyses of V0v Spinal Interneurons and Identification of Transcriptional Regulators Downstream of Evx1 and Evx2 in These Cells. [scRNA Seq]	GSE240239	Transcriptome Analysis	Background: V0v spinal interneurons are highly conserved glutamatergic commissural neurons that function in locomotor circuits. We have previously shown that Evx1 and Evx2 are required to specify the neurotransmitter phenotype of these cells. However we still know very little about the gene regulatory networks that act downstream of these transcription factors in V0v cells. Methods: To identify candidate members of V0v gene regulatory networks we FAC sorted WT and evx1;evx2 double mutant zebrafish V0v spinal interneurons and expression profiled them using microarrays and scRNA seq. We also used in situ hybridization to compare expression of a subset of candidate genes in evx1;evx2 mutants and wild type siblings. Results: Our data reveal two molecularly distinct subtypes of V0v spinal interneurons at 48 h and suggest that by this stage of development evx1;evx2 double mutant cells transfate into either inhibitory spinal interneurons or motoneurons. Our results also identify 25 transcriptional regulator genes that require Evx1/2 for their expression in V0v interneurons plus a further 11 transcriptional regulator genes that are repressed in V0v interneurons by Evx1/2. Two of the latter genes are hmx2 and hmx3a. Intriguingly we show that Hmx2/3a repress dI2 interneuronal expression of skor1a and nefma two genes that require Evx1/2 for their expression in V0v interneurons. This suggests that Evx1/2 might regulate skor1a and nefma expression in V0v interneurons by repressing Hmx2/3a expression. Conclusions: This study identifies two molecularly distinct subsets of V0v spinal interneurons as well as multiple transcriptional regulators that are strong candidates for acting downstream of Evx1/2 to specify the essential functional characteristics of V0v interneurons. Our data further suggest that in the absence of both Evx1 and Evx2 V0v spinal interneurons initially change their neurotransmitter phenotypes from excitatory to inhibitory and then later start to express markers of distinct types of inhibitory s…	parent bioproject:PRJNA1003022	pubmed:38017520		MUTSU2 OtherGFP G8	GSM7688795		source name:Spinal Cord\|tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1\|geo loc name:missing\|collection date:missing	MUTSU2 OtherGFP G8	We performed demultiplexing and counts analysis as per the manufacturer’s instructions using Cell Ranger v4.0.0 software https://www.10xgenomics.com We analyzed the data using Partek Flow Genomic Analysis Software https://www.partek.com/partek flow/. Multiplets were removed by filtering out cells with >12 000 counts and >2 500 detected genes. Sick and/or “leaky” cells were removed by filtering out cells with <500 detected genes and >6% mitochondrial transcripts. We normalized the data using a counts per million CPM algorithm and applied a logarithmic transformation to improve data visualization. The outcome of normalisation was assessed by principal components analysis PCA graph based clustering and Uniform Manifold Approximation and Projection UMAP plotting using the NN Descent method of nearest neighbour type calculation and Euclidean distance metrics. We manually inspected UMAP plots to assess clustering quality based on expression of known V0v spinal interneuron markers. We then fine tuned the clustering by manually deducing and extrapolating cell fate assignments by comparing expression profiles of 48 hpf single cell clusters with the molecular phenotypes of V0v spinal interneurons in 24 hpf and 30 hpf wild type evx1i232 and evx2sa140 single mutant and evx1i232;evx2sa140 double mutant embryos. To perform differential expression analysis in Partek Flow we used the statistically robust Hurdle Model two part model with default parameters. We initially analyzed the two libraries separately. We then combined the data from these two libraries using the Counts Aggregation pipeline in Cell Ranger v4.0.0 and reanalyzed the data as described above. Assembly: Lawson Lab zebrafish transcriptome V4.3.2 https://www.umassmed.edu/lawson lab/reagents/zebrafish transcriptome/ Supplementary files format and content: Tab separated and matrix files.	Spinal Cord		Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz’s L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…	Embryos obtained from crossing heterozygous evx1i232/+;evx2sa140/+;Tgevx1:EGFPSU2 parents were raised at 28.5oC until they reached the desired developmental stage of 48 hpf as confirmed by analysis of morphological criteria including head trunk angle head and eye size.	tissue:Spinal Cord\|cell line:Tgevx1:EGFPSU2\|cell type:V0v spinal interneurons\|genotype:evx1i232/+;evx2sa140/+ incross\|treatment:N1	GSM7688795	GSM7688795: MUTSU2 OtherGFP G8; Danio rerio; RNA Seq	GSM7688795 r1	GSM7688795	1	Embryos were screened for fluorescence from 30 hpf onwards using a fluorescent dissecting microscope. Only EGFP positive embryos were used for dissections and FACS at 48 hpf. Embryos were deyolked dissected and dissociated as described in GSE145916 with the following modifications: Trunk tissue was dissected anteriorly at the boundary between the hindbrain and spinal cord and posteriorly immediately above the end of the yolk extension. To ensure complete dissociation of trunk tissue with the Papain Dissociation System Worthington Biochemical Corporation LK003150 trunks were incubated in 1 ml Papain/DNase mix with gentle rocking at 28.5oC for 40 minutes. The digested tissue was then allowed to settle for 10 seconds before the Papain/DNase mix was carefully decanted until approximately 500 µl remained. Immediately post homogenising the digested tissue mixture with a sterile p200 tip we passed each sample through a 40 µm Flowmi cell strainer Merck BAH136800040 into a sterile microcentrifuge tube. post Papain inactivation samples were resuspended in 1 ml Leibovitz's L 15 medium ThermoFisher Scientific 21083027 + 0.5% FBS and stored on ice. Immediately before FACS DAPI Merck D9542 and Draq5 BioLegend 424101 were added at a final concentration of 5 µg/ml and 5 µM respectively. FACS was performed using a Becton Dickinson FACS Aria III Cell Sorter at the SUNY Upstate Medical University Research Flow Core using the parameters described by Cerda et al. 2008 with the following modifications. Ice cold samples were filtered through 35 µm mesh strainers in to 5 ml round bottomed polystyrene tubes Corning Falcon 352235. All FAC sorting and collection steps were performed at +4oC using a 100 µm nozzle and 20 psi sort pressure. Successive doublet exclusion gates forward scatter height x forward scatter width followed by side scatter height x side scatter width were used to finesse capture of real single cells. Accurate live/dead filtering was performed by selecting for DAPI negative sick cells are DAPI permeant…		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP453891		loader:fastq load.py	MUTSU2_OtherGFP-G8-3_S1_L001_R2_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L001_R1_001.fastq.gz MUTSU2_OtherGFP-G8-3_S1_L001_I1_001.fastq.gz	fastq fastq fastq	8184961024.0	64448512.0	GSM7688795 r9	0:8 1:28 2:91	A:1748819280;C:1187802244;G:1360994184;T:1562975470;N:4223414	8	28	91	1748819280	1187802244	1360994184	1562975470	4223414	SRX21286762	SRS18536875	SRA1688438	Lewis Lab, Biology, Syracuse University	Lewis Lab, Biology, Syracuse University	1	0.8493		0.21072		0.80967		0.52593		91		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-08-07	Multi-stage	Embryo	Spinal Cord	Nervous System
60587	60587	SRR12424278	SRX8920145	SRS7176693	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 48hpf	GSM4718658		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718658	GSM4718658: scRNAseq olig2 eGFP 48hpf; Danio rerio; RNA Seq	GSM4718658		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718658	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			olig2_egfp_48hpf_1_S41_L001_R1_001.fastq.gz olig2_egfp_48hpf_1_S41_L001_R2_001.fastq.gz	fastq fastq	18778651430.0	62180965.0	GSM4718658 r1	0:151 1:151	A:6542554044;C:3515052559;G:3489753648;T:5231107020;N:184159	151	151		6542554044	3515052559	3489753648	5231107020	184159	SRX8920145	SRS7176693	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.90055	0.0	0.21162	1.0	0.77776		0.53718	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Hatching	Embryo	Spinal Cord	Nervous System
60588	60588	SRR12424279	SRX8920145	SRS7176693	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 48hpf	GSM4718658		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718658	GSM4718658: scRNAseq olig2 eGFP 48hpf; Danio rerio; RNA Seq	GSM4718658		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718658	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			olig2_egfp_48hpf_2_S42_L001_R1_001.fastq.gz olig2_egfp_48hpf_2_S42_L001_R2_001.fastq.gz	fastq fastq	22242782898.0	73651599.0	GSM4718658 r2	0:151 1:151	A:7753674261;C:4160150132;G:4127581173;T:6201157562;N:219770	151	151		7753674261	4160150132	4127581173	6201157562	219770	SRX8920145	SRS7176693	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.90097	0.0	0.2127	1.0	0.77966		0.54192	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Hatching	Embryo	Spinal Cord	Nervous System
60589	60589	SRR12424280	SRX8920145	SRS7176693	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 48hpf	GSM4718658		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718658	GSM4718658: scRNAseq olig2 eGFP 48hpf; Danio rerio; RNA Seq	GSM4718658		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718658	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			olig2_egfp_48hpf_3_S43_L001_R2_001.fastq.gz olig2_egfp_48hpf_3_S43_L001_R1_001.fastq.gz	fastq fastq	25591190986.0	84739043.0	GSM4718658 r3	0:151 1:151	A:8917343424;C:4797392797;G:4757647263;T:7118554564;N:252938	151	151		8917343424	4797392797	4757647263	7118554564	252938	SRX8920145	SRS7176693	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.90089	0.0	0.2095	1.0	0.78078		0.53755	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Hatching	Embryo	Spinal Cord	Nervous System
60590	60590	SRR12424281	SRX8920145	SRS7176693	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 48hpf	GSM4718658		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718658	GSM4718658: scRNAseq olig2 eGFP 48hpf; Danio rerio; RNA Seq	GSM4718658		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718658	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			olig2_egfp_48hpf_4_S44_L001_R1_001.fastq.gz olig2_egfp_48hpf_4_S44_L001_R2_001.fastq.gz	fastq fastq	25546281170.0	84590335.0	GSM4718658 r4	0:151 1:151	A:8905009196;C:4774524700;G:4734395060;T:7132103501;N:248713	151	151		8905009196	4774524700	4734395060	7132103501	248713	SRX8920145	SRS7176693	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.90169	0.0	0.21158	1.0	0.77697		0.54006	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Hatching	Embryo	Spinal Cord	Nervous System
60591	60591	SRR12424274	SRX8920144	SRS7176692	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 36hpf	GSM4718657		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718657	GSM4718657: scRNAseq olig2 eGFP 36hpf; Danio rerio; RNA Seq	GSM4718657		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718657	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_36hfp_1_S69_L001_R1_001.fastq.gz Olig2_eGFP_36hfp_1_S69_L001_R2_001.fastq.gz	fastq fastq	13366864582.0	44261141.0	GSM4718657 r1	0:151 1:151	A:3721647646;C:2385939960;G:3494552257;T:3764514262;N:210457	151	151		3721647646	2385939960	3494552257	3764514262	210457	SRX8920144	SRS7176692	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.85449	0.0	0.08264	1.0	0.84086		0.50156	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60592	60592	SRR12424275	SRX8920144	SRS7176692	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 36hpf	GSM4718657		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718657	GSM4718657: scRNAseq olig2 eGFP 36hpf; Danio rerio; RNA Seq	GSM4718657		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718657	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_36hfp_2_S70_L001_R1_001.fastq.gz Olig2_eGFP_36hfp_2_S70_L001_R2_001.fastq.gz	fastq fastq	10606211914.0	35119907.0	GSM4718657 r2	0:151 1:151	A:2974656607;C:1869869441;G:2750621038;T:3010898673;N:166155	151	151		2974656607	1869869441	2750621038	3010898673	166155	SRX8920144	SRS7176692	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.84822	0.0	0.08456	1.0	0.83719		0.51503	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60593	60593	SRR12424276	SRX8920144	SRS7176692	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 36hpf	GSM4718657		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718657	GSM4718657: scRNAseq olig2 eGFP 36hpf; Danio rerio; RNA Seq	GSM4718657		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718657	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_36hfp_3_S71_L001_R1_001.fastq.gz Olig2_eGFP_36hfp_3_S71_L001_R2_001.fastq.gz	fastq fastq	15614508642.0	51703671.0	GSM4718657 r3	0:151 1:151	A:4330312984;C:2826855730;G:4093245712;T:4363847925;N:246291	151	151		4330312984	2826855730	4093245712	4363847925	246291	SRX8920144	SRS7176692	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.8543	0.0	0.08219	1.0	0.83727		0.51892	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60594	60594	SRR12424277	SRX8920144	SRS7176692	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 36hpf	GSM4718657		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718657	GSM4718657: scRNAseq olig2 eGFP 36hpf; Danio rerio; RNA Seq	GSM4718657		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718657	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_36hfp_4_S72_L001_R1_001.fastq.gz Olig2_eGFP_36hfp_4_S72_L001_R2_001.fastq.gz	fastq fastq	10707787198.0	35456249.0	GSM4718657 r4	0:151 1:151	A:2981466900;C:1909696186;G:2797297473;T:3019156550;N:170089	151	151		2981466900	1909696186	2797297473	3019156550	170089	SRX8920144	SRS7176692	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.85466	0.0	0.08381	1.0	0.83562		0.51035	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60595	60595	SRR12424270	SRX8920143	SRS7176691	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 24hpf	GSM4718656		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718656	GSM4718656: scRNAseq olig2 eGFP 24hpf; Danio rerio; RNA Seq	GSM4718656		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718656	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_24hfp_1_S10_L001_R1_001.fastq.gz Olig2_eGFP_24hfp_1_S10_L001_R2_001.fastq.gz	fastq fastq	21279021170.0	70460335.0	GSM4718656 r1	0:151 1:151	A:5842145219;C:3712477572;G:5933108587;T:5790922441;N:367351	151	151		5842145219	3712477572	5933108587	5790922441	367351	SRX8920143	SRS7176691	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.83071	0.0	0.06828	1.0	0.8406		0.5078	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60596	60596	SRR12424271	SRX8920143	SRS7176691	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 24hpf	GSM4718656		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718656	GSM4718656: scRNAseq olig2 eGFP 24hpf; Danio rerio; RNA Seq	GSM4718656		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718656	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_24hfp_2_S20_L001_R1_001.fastq.gz Olig2_eGFP_24hfp_2_S20_L001_R2_001.fastq.gz	fastq fastq	22414094210.0	74218855.0	GSM4718656 r2	0:151 1:151	A:6145677154;C:3904876630;G:6254105391;T:6109049174;N:385861	151	151		6145677154	3904876630	6254105391	6109049174	385861	SRX8920143	SRS7176691	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.83148	0.0	0.06914	1.0	0.84033		0.49666	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60597	60597	SRR12424272	SRX8920143	SRS7176691	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 24hpf	GSM4718656		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718656	GSM4718656: scRNAseq olig2 eGFP 24hpf; Danio rerio; RNA Seq	GSM4718656		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718656	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_24hfp_3_S30_L001_R1_001.fastq.gz Olig2_eGFP_24hfp_3_S30_L001_R2_001.fastq.gz	fastq fastq	16406166006.0	54325053.0	GSM4718656 r3	0:151 1:151	A:4513103872;C:2865727260;G:4569430659;T:4457621166;N:283049	151	151		4513103872	2865727260	4569430659	4457621166	283049	SRX8920143	SRS7176691	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.82928	0.0	0.06878	1.0	0.83989		0.4916	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
60598	60598	SRR12424273	SRX8920143	SRS7176691	SRP276929	PRJNA656271	Single cell RNA seq analysis of pMN neural progenitors from zebrafish	GSE155988	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:32680935		scRNAseq olig2 eGFP 24hpf	GSM4718656		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf	Raw sequencing reads were demultiplexed mapped to the zebrafish reference genome and summarized into gene expression matrices using CellRanger version 3.0.1 Count matrices were further filtered in Seurat 3.1.0 https://satijalab.org/seurat/ to remove cell barcodes with fewer than 250 detectable genes more than 5% of UMIs derived from mitochondrial genes or more than 50 000 UMIs to exclude putative doublets Standard Seurat normalization and PCA was run using the 1 291 most variable genes Applied Harmony https://doi.org/10.1038/s41592 019 0619 0 alignment theta = 2 to correct for inter sample variation chose 20 Harmony dimensions and applied Seurat UMAP reduction Genome build: GRCz11 Supplementary files format and content: gzipped csv file of matrix of normalized gene expression by cell for merged 3 scRNAseq samples Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM4718656	GSM4718656: scRNAseq olig2 eGFP 24hpf; Danio rerio; RNA Seq	GSM4718656		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM4718656	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP276929			Olig2_eGFP_24hfp_4_S40_L001_R1_001.fastq.gz Olig2_eGFP_24hfp_4_S40_L001_R2_001.fastq.gz	fastq fastq	17382967792.0	57559496.0	GSM4718656 r4	0:151 1:151	A:4767137063;C:3044788720;G:4846983492;T:4723757048;N:301469	151	151		4767137063	3044788720	4846983492	4723757048	301469	SRX8920143	SRS7176691	SRA1111061	GEO	University of Colorado School of Medicine	2	0.0	0.83084	0.0	0.06779	1.0	0.83948		0.49689	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2020-08-10	Pharyngula	Embryo	Spinal Cord	Nervous System
62942	62942	SRR13447021	SRX9860257	SRS8040688	SRP301983	PRJNA692494	Localized EMT reprograms glial progenitors to promote spinal cord repair [single cell RNA seq]	GSE164944	Other	Single nuclear RNA sequencing was performed on spinal cord tissues from Tggfap:EGFP zebrafish at 1 xxx post injury using the 10x Genomics platform. Overall design: Single nuclear RNA sequencing of spinal cord tissue.	parent bioproject:PRJNA692492	pubmed:33609461		single cell injured spinal cord	GSM5023610		tissue:5 xxx post injury spinal cord nuclear\|tissues:spinal cord\|rna population:nuclear\|time point:5 xxx post injury	single cell injured spinal cord	Aligned reads were analyzed using the Seurat package v3.1.4 within R v3.6.3. Cells that have unique number of genes between 200 to 4000 and a mitochondrial gene percentage less than five were filtered. The “SCTransform” function was used for normalization scaling and finding variable features. SCTransform returned 3000 highly variable features for downstream analysis. Significant principal components were determined using “ElbowPlot” function. Forty principal components were selected to create a Shared Nearest Neighbour SNN graph using the “FindNeighbours” function. Clustering was performed by the Louvain algorithm using the function “FindClusters” and the resolution parameter was set to one. Uniform Manifold Approximation and Reduction UMAP was used for non linear dimensional reduction of the first 40 principle components and to visualize the data in two dimensional space “RunUMAP” function Differential gene expression for individual cluster was identified using Wilcoxon rank sum tests in the “FindAllMarkers” function. Supplementary files format and content: RDS file	5 xxx post injury spinal cord nuclear	Adult zebrafish were subjected to complete spinal cord transection.	gfap:EGFP transgenic animals were subjected to SCI and 2 mm SC tissue sections spanning the lesion site were collected at 1 wpi. Nuclei were isolated from 45 pooled SC tissues. For single nuclear library preparation on the 10x Genomics platform the Chromium Single Cell 3′ GEM Library and Gel Bead Kit v3 cat# 1000092 and the Chromium Chip B Single Cell Kit cat# 1000074 were used according to the manufacturer’s instructions in the Chromium Single Cell 3′ Reagents Kits V3 User Guide. The resulting cDNA libraries were quantified on an Agilent Tapestation and sequenced on Illumina HiSeq 3000.		tissues:spinal cord\|rna population:nuclear\|time point:5 xxx post injury	GSM5023610	GSM5023610: single cell injured spinal cord; Danio rerio; RNA Seq	GSM5023610		1	gfap:EGFP transgenic animals were subjected to SCI and 2 mm SC tissue sections spanning the lesion site were collected at 1 wpi. Nuclei were isolated from 45 pooled SC tissues. For single nuclear library preparation on the 10x Genomics platform the Chromium Single Cell 3′ GEM Library and Gel Bead Kit v3 cat# 1000092 and the Chromium Chip B Single Cell Kit cat# 1000074 were used according to the manufacturer's instructions in the Chromium Single Cell 3′ Reagents Kits V3 User Guide. The resulting cDNA libraries were quantified on an Agilent Tapestation and sequenced on Illumina HiSeq 3000.	GEO Accession:GSM5023610	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina HiSeq 3000	SRP301983		loader:fastq load.py\|options: platform=Illumina readTypes=TTB read1PairFiles=ZB nuc S1 L007 I1 001.fastq.gz read2PairFiles=ZB nuc S1 L007 R1 001.fastq.gz read3PairFiles=ZB nuc S1 L007 R2 001.fastq.gz	ZB_nuc_S1_L007_R2_001.fastq.gz ZB_nuc_S1_L007_R1_001.fastq.gz ZB_nuc_S1_L007_I1_001.fastq.gz	fastq fastq fastq	134333186988.0	436146711.0	GSM5023610 r1	0:8 1:150 2:150	A:34573306207;C:23174725516;G:24790836286;T:51771968988;N:22349991	8	150	150	34573306207	23174725516	24790836286	51771968988	22349991	SRX9860257	SRS8040688	SRA1184553	GEO	Developmental Biology, Washington University	1	0.72186		0.41754		0.80551		0.54979		150		B		usable mapping rate	illumina	hiseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-01-15	Adult	Adult	Spinal Cord	Nervous System
64205	64205	SRR14326888	SRX10681834	SRS8775213	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 48hpf [reanalysis]	GSM5266260		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266260	GSM5266260: scRNAseq olig2 eGFP 48hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266260		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266260	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			olig2_egfp_48hpf_1_S41_L001_R1_001.fastq olig2_egfp_48hpf_1_S41_L001_R2_001.fastq	fastq fastq	18778651430.0	62180965.0	GSM5266260 r1	0:151 1:151	A:6542554044;C:3515052559;G:3489753648;T:5231107020;N:184159	151	151		6542554044	3515052559	3489753648	5231107020	184159	SRX10681834	SRS8775213	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.90056	0.0	0.21167	1.0	0.7781		0.53738	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Hatching	Embryo	Spinal Cord	Nervous System
64206	64206	SRR14326889	SRX10681834	SRS8775213	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 48hpf [reanalysis]	GSM5266260		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266260	GSM5266260: scRNAseq olig2 eGFP 48hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266260		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266260	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			olig2_egfp_48hpf_2_S42_L001_R1_001.fastq olig2_egfp_48hpf_2_S42_L001_R2_001.fastq	fastq fastq	22242782898.0	73651599.0	GSM5266260 r2	0:151 1:151	A:7753674261;C:4160150132;G:4127581173;T:6201157562;N:219770	151	151		7753674261	4160150132	4127581173	6201157562	219770	SRX10681834	SRS8775213	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.90096	0.0	0.21256	1.0	0.77928		0.54059	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Hatching	Embryo	Spinal Cord	Nervous System
64207	64207	SRR14326890	SRX10681834	SRS8775213	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 48hpf [reanalysis]	GSM5266260		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266260	GSM5266260: scRNAseq olig2 eGFP 48hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266260		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266260	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			olig2_egfp_48hpf_3_S43_L001_R2_001.fastq olig2_egfp_48hpf_3_S43_L001_R1_001.fastq	fastq fastq	25591190986.0	84739043.0	GSM5266260 r3	0:151 1:151	A:8917343424;C:4797392797;G:4757647263;T:7118554564;N:252938	151	151		8917343424	4797392797	4757647263	7118554564	252938	SRX10681834	SRS8775213	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.90089	0.0	0.20934	1.0	0.78086		0.53725	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Hatching	Embryo	Spinal Cord	Nervous System
64208	64208	SRR14326891	SRX10681834	SRS8775213	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 48hpf [reanalysis]	GSM5266260		tissue:spinal cord cells\|developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 48hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:48 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266260	GSM5266260: scRNAseq olig2 eGFP 48hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266260		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266260	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			olig2_egfp_48hpf_4_S44_L001_R1_001.fastq olig2_egfp_48hpf_4_S44_L001_R2_001.fastq	fastq fastq	25546281170.0	84590335.0	GSM5266260 r4	0:151 1:151	A:8905009196;C:4774524700;G:4734395060;T:7132103501;N:248713	151	151		8905009196	4774524700	4734395060	7132103501	248713	SRX10681834	SRS8775213	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.90171	0.0	0.21154	1.0	0.77674		0.53902	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Hatching	Embryo	Spinal Cord	Nervous System
64209	64209	SRR14326884	SRX10681833	SRS8775212	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 36hpf [reanalysis]	GSM5266259		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266259	GSM5266259: scRNAseq olig2 eGFP 36hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266259		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266259	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_36hfp_1_S69_L001_R1_001.fastq Olig2_eGFP_36hfp_1_S69_L001_R2_001.fastq	fastq fastq	13366864582.0	44261141.0	GSM5266259 r1	0:151 1:151	A:3721647646;C:2385939960;G:3494552257;T:3764514262;N:210457	151	151		3721647646	2385939960	3494552257	3764514262	210457	SRX10681833	SRS8775212	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.85451	0.0	0.08269	1.0	0.84094		0.5016	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64210	64210	SRR14326885	SRX10681833	SRS8775212	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 36hpf [reanalysis]	GSM5266259		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266259	GSM5266259: scRNAseq olig2 eGFP 36hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266259		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266259	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_36hfp_2_S70_L001_R1_001.fastq Olig2_eGFP_36hfp_2_S70_L001_R2_001.fastq	fastq fastq	10606211914.0	35119907.0	GSM5266259 r2	0:151 1:151	A:2974656607;C:1869869441;G:2750621038;T:3010898673;N:166155	151	151		2974656607	1869869441	2750621038	3010898673	166155	SRX10681833	SRS8775212	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.84821	0.0	0.08459	1.0	0.83731		0.51513	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64211	64211	SRR14326886	SRX10681833	SRS8775212	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 36hpf [reanalysis]	GSM5266259		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266259	GSM5266259: scRNAseq olig2 eGFP 36hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266259		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266259	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_36hfp_3_S71_L001_R1_001.fastq Olig2_eGFP_36hfp_3_S71_L001_R2_001.fastq	fastq fastq	15614508642.0	51703671.0	GSM5266259 r3	0:151 1:151	A:4330312984;C:2826855730;G:4093245712;T:4363847925;N:246291	151	151		4330312984	2826855730	4093245712	4363847925	246291	SRX10681833	SRS8775212	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.85427	0.0	0.08218	1.0	0.83713		0.51889	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64212	64212	SRR14326887	SRX10681833	SRS8775212	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 36hpf [reanalysis]	GSM5266259		tissue:spinal cord cells\|developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 36hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:36 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266259	GSM5266259: scRNAseq olig2 eGFP 36hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266259		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266259	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_36hfp_4_S72_L001_R1_001.fastq Olig2_eGFP_36hfp_4_S72_L001_R2_001.fastq	fastq fastq	10707787198.0	35456249.0	GSM5266259 r4	0:151 1:151	A:2981466900;C:1909696186;G:2797297473;T:3019156550;N:170089	151	151		2981466900	1909696186	2797297473	3019156550	170089	SRX10681833	SRS8775212	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.85465	0.0	0.08381	1.0	0.8355		0.51049	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64213	64213	SRR14326880	SRX10681832	SRS8775211	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 24hpf [reanalysis]	GSM5266258		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266258	GSM5266258: scRNAseq olig2 eGFP 24hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266258		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266258	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_24hfp_1_S10_L001_R1_001.fastq Olig2_eGFP_24hfp_1_S10_L001_R2_001.fastq	fastq fastq	21279021170.0	70460335.0	GSM5266258 r1	0:151 1:151	A:5842145219;C:3712477572;G:5933108587;T:5790922441;N:367351	151	151		5842145219	3712477572	5933108587	5790922441	367351	SRX10681832	SRS8775211	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.83071	0.0	0.06834	1.0	0.8407		0.50743	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64214	64214	SRR14326881	SRX10681832	SRS8775211	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 24hpf [reanalysis]	GSM5266258		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266258	GSM5266258: scRNAseq olig2 eGFP 24hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266258		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266258	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_24hfp_2_S20_L001_R1_001.fastq Olig2_eGFP_24hfp_2_S20_L001_R2_001.fastq	fastq fastq	22414094210.0	74218855.0	GSM5266258 r2	0:151 1:151	A:6145677154;C:3904876630;G:6254105391;T:6109049174;N:385861	151	151		6145677154	3904876630	6254105391	6109049174	385861	SRX10681832	SRS8775211	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.83142	0.0	0.06899	1.0	0.84029		0.49666	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64215	64215	SRR14326882	SRX10681832	SRS8775211	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 24hpf [reanalysis]	GSM5266258		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266258	GSM5266258: scRNAseq olig2 eGFP 24hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266258		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266258	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_24hfp_3_S30_L001_R1_001.fastq Olig2_eGFP_24hfp_3_S30_L001_R2_001.fastq	fastq fastq	16406166006.0	54325053.0	GSM5266258 r3	0:151 1:151	A:4513103872;C:2865727260;G:4569430659;T:4457621166;N:283049	151	151		4513103872	2865727260	4569430659	4457621166	283049	SRX10681832	SRS8775211	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.8293	0.0	0.06878	1.0	0.84017		0.49152	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
64216	64216	SRR14326883	SRX10681832	SRS8775211	SRP316409	PRJNA725224	Single cell RNA seq analysis of pMN neural progenitors from zebrafish [Reanalysis of GSE155988 data set]	GSE173350	Transcriptome Analysis	The goal of this study was to identify distinct cell populations that arise from ventral spinal cord pMN progenitors. To do so we sorted fluorescently marked pMN cells obtained from Tgolig2:EGFP zebrafish embryos at 24 36 hpf and 48 hpf and performed 10X Chromium single cell RNA seq. Overall design: Three samples obtained from wild type zebrafish embryos at three developmental timepoints were analyzed.		pubmed:34303700		scRNAseq olig2 eGFP 24hpf [reanalysis]	GSM5266258		tissue:spinal cord cells\|developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	scRNAseq olig2 eGFP 24hpf [reanalysis]	Generated cell feature counts for each single cell RNAseq library using cellranger count transcriptome=GRCz11 Each single cell RNAseq sample was converted to a Seurat object using the Seurat Read 10X CreateSeuratObject functions 3 separate Seurat objects filtered individual Seurat objects to select cells with nGene >250 and percent.mitochondrial <5% and nFeature RNA <50000 hpf 24 hpf and 48 hpf samples or <30000 hpf 36 hpf Processed and filtered Seurat objects using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction 'pca' dims = 1:40 FindClustersresolution= 0.9 hpf 48 hpf; 1 hpf 36 hpf; 1.2 hpf 24 hpf dims = 1:40 and RunUMAPdims=1:40 Clusters with low nFeatures and high percent.mitochondrial reads were removed as dying cells in 24 hpf and 36 hpf datasets Individual processed Seurat objects from each sample were intergrated using the FindIntergrationAnchorsdims 1:40 and IntergrateDatadims = 1:40 Seurat functions Processed and filtered intergrated Seurat object using Seurat NormalizeData FindVariableFeaturesselection.method = 'vst' nfeatures = 2000 ScaleData vars.to.regress=c'nCount RNA' 'percent.mito' 'CC.difference' RunPCAnpcs=100 RunUMAPdims = 1:40 FindNeighborsreduction = 'pca' dims = 1:40 FindClustersresolution= 1.2 dims = 1:40 and RunUMAPdims=1:40 Genome build: GRCz11 Supplementary files format and content: gzipped rds file of intergrated Seurat object Supplementary files format and content: gzipped csv file of metadata for cells	spinal cord cells	No treatment was applied to the samples.	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	Embryos were raised at 28.5°C in E3 media 5 mM NaCl 0.17 mM KCl 0.33 mM CaCl2 0.33 mM MgSO4 at pH 7.4 with sodium bicarbonate sorted for good health and staged accordingly to developmental morphological features and hpf	developmental stage:24 hpf ZFIN ID: ZDB ALT 041129 8	GSM5266258	GSM5266258: scRNAseq olig2 eGFP 24hpf [reanalysis]; Danio rerio; RNA Seq	GSM5266258		1	24 36 and 48 hpf Tgolig2:EGFP euthanized embryos were collected in 1.7 ml microcentrifuge tubes and deyolked in 100 μl of pre chilled Ca free Ringers solution 116 mM NaCl 2.6 mM KCl 5 mM HEPES pH 7.0 on ice. Embryos were pipetted intermittently with a p200 micropipettor for 15 minutes and left for 5 min. 500 μl of protease solution 10 mg/ml BI protease 125 U/ml DNase 2.5 mM EDTA 1X PBS was added to microcentrifuge tubes on ice for 15 min and embryos were homogenized every 3 min with a p100 micropipettor for 15 min. 200 μl of STOP solution 30% FBS 0.8 mM CaCl2 1X PBS was then mixed into the tubes. Samples were then spun down at 400g for 5 min at 4°C and supernatant was removed. On ice 1 ml of chilled suspension media 1% FBS 0.8 mM CaCl2 50 U/ml Penicillin 0.05 mg/ml Streptomycin was added to samples and then spun down again at 400g for 5 min at 4°C. Supernatant was removed and 400 μl of chilled suspension media was added and solution was filtered through a 35 μm strainer into a collection tube. Cells were FAC sorted to distinguish EGFP+ cells using a MoFlo XDP100 cell sorter at the CU SOM Cancer Center Flow Cytometry Shared Resource and collected in 1.7 ml FBS coated microcentrifuge tubes in 200 μl of 1X PBS. The Chromium Box from 10X Genomics was used to capture cells using Chromium Single Cell three prime Reagent Kit part no. PN 1000075. Libraries were sequenced on the Illumina NovaSEQ6000 Instrument.	GEO Accession:GSM5266258	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP316409			Olig2_eGFP_24hfp_4_S40_L001_R1_001.fastq Olig2_eGFP_24hfp_4_S40_L001_R2_001.fastq	fastq fastq	17382967792.0	57559496.0	GSM5266258 r4	0:151 1:151	A:4767137063;C:3044788720;G:4846983492;T:4723757048;N:301469	151	151		4767137063	3044788720	4846983492	4723757048	301469	SRX10681832	SRS8775211	SRA1223938	GEO	Bruce Appel, Pediatrics, Section of Developmental Biology, University of Colorado Anschutz Medical Campus	2	0.0	0.83086	0.0	0.0679	1.0	0.83944		0.4968	151	151	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2021-04-26	Pharyngula	Embryo	Spinal Cord	Nervous System
74357	74357	SRR23690175	SRX19552676	SRS16937434	SRP425371	PRJNA940435	sox1a:eGFP transgenic line and single cell transcriptomics reveal the origin of zebrafish intraspinal serotonergic neurons	GSE226494	Transcriptome Analysis	The Sox family of transcription factors plays a crucial role in the development of the vertebrate nervous system. In the zebrafish embryo sox1 genes are expressed in neural progenitor cells and neurons of the ventral spinal cord. We recently reported that the loss of function of sox1a and sox1b results in a significant decrease in a subtype of V2 neurons called V2s in zebrafish. Here a single cell RNA sequencing scRNA Seq approach was used to analyse the transcriptome of sox1a lineage progenitors and neurons in the zebrafish spinal cord at four different time points during the development of an earlier established Tgsox1a:EGFP line. In addition to the sox1a expressing neurons described previously we found that this gene is also expressed in late developing intraspinal serotonin neurons ISNs. Developmental trajectory analysis of single cell data and morpholino knockdown of lateral floor plate LFP cells expressing the nkx2.9 gene suggest that ISNs are generated from LFP progenitor cells. Pharmacological inhibition of the Notch signalling pathway demonstrates the necessity of this pathway for the development of LFP progenitor cells into ISN populations. Our results confirm that the zebrafish LFP is a progenitor domain from which ISN neurons are generated in addition to the previously described KA" and V3. Overall design: The trunk and tail of Tgsox1a:eGFP lines were and dissected at 1 2 dpf 3 dpf and 5 dpf. The tissue was dissociated and GFP positive cells were sorted by FACS as described Cosacak et al. 2019. The single cell encapsulation cDNA synthesis by 10X Genomics. The reads were aligned to zebrafish genom GRChZ 11 and ensemble version 105 was used for gene annotation and assigning reads to genes. Genomics Kit and Sequencing done by Illumina.		pubmed:37529101		sox1a 5dpf	GSM7077910		source name:spinal cord\|tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:5dpf\|geo loc name:missing\|collection date:missing	sox1a 5dpf	The demultiplexing barcoded processing gene counting and aggregation were made using the 10x genomics' cellranger software version 6.1.2 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/output/metrics The fastq files were aligned to zebrafish genome GRCz11 and ensemble transcripts from Ensembl Release 104 by using STAR. The BAM files were as input for Cell Ranger 10X genmoics to generate processed data files that include gene names row names and cell names column names and counts. Further analysis done by using Seurat package in R. Assembly: GRCz11 Supplementary files format and content: filtered bc matrix outputs from Cell Ranger	spinal cord	No treatment done.	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics	Zebrafish were kept in the re circulating system on a 14/10 h light/dark cycle pH 7.5 at 28 °C ±1 °C in groups of 20 animals per 2.8 L.	tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:5dpf	GSM7077910	GSM7077910: sox1a 5dpf; Danio rerio; RNA Seq	GSM7077910 r1	GSM7077910	1	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP425371		loader:fastq load.py	L100349_Track-139094_R1.fastq.gz L100349_Track-139094_R2.fastq.gz	fastq fastq	78287518950.0	387561975.0	GSM7077910 r1	0:101 1:101	A:24402003655;C:12853163240;G:13012950072;T:28016227491;N:3174492	101	101		24402003655	12853163240	13012950072	28016227491	3174492	SRX19552676	SRS16937434	SRA1601794	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	2	0.0	0.8342	0.0	0.22003	1.0	0.75797		0.52045	101	101	T	B	mate1 technical by mapping diff	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	Germany	2023-03-02	Larval	Larval	Spinal Cord	Nervous System
74358	74358	SRR23690176	SRX19552675	SRS16937433	SRP425371	PRJNA940435	sox1a:eGFP transgenic line and single cell transcriptomics reveal the origin of zebrafish intraspinal serotonergic neurons	GSE226494	Transcriptome Analysis	The Sox family of transcription factors plays a crucial role in the development of the vertebrate nervous system. In the zebrafish embryo sox1 genes are expressed in neural progenitor cells and neurons of the ventral spinal cord. We recently reported that the loss of function of sox1a and sox1b results in a significant decrease in a subtype of V2 neurons called V2s in zebrafish. Here a single cell RNA sequencing scRNA Seq approach was used to analyse the transcriptome of sox1a lineage progenitors and neurons in the zebrafish spinal cord at four different time points during the development of an earlier established Tgsox1a:EGFP line. In addition to the sox1a expressing neurons described previously we found that this gene is also expressed in late developing intraspinal serotonin neurons ISNs. Developmental trajectory analysis of single cell data and morpholino knockdown of lateral floor plate LFP cells expressing the nkx2.9 gene suggest that ISNs are generated from LFP progenitor cells. Pharmacological inhibition of the Notch signalling pathway demonstrates the necessity of this pathway for the development of LFP progenitor cells into ISN populations. Our results confirm that the zebrafish LFP is a progenitor domain from which ISN neurons are generated in addition to the previously described KA" and V3. Overall design: The trunk and tail of Tgsox1a:eGFP lines were and dissected at 1 2 dpf 3 dpf and 5 dpf. The tissue was dissociated and GFP positive cells were sorted by FACS as described Cosacak et al. 2019. The single cell encapsulation cDNA synthesis by 10X Genomics. The reads were aligned to zebrafish genom GRChZ 11 and ensemble version 105 was used for gene annotation and assigning reads to genes. Genomics Kit and Sequencing done by Illumina.		pubmed:37529101		sox1a 3dpf	GSM7077909		source name:spinal cord\|tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:3dpf\|geo loc name:missing\|collection date:missing	sox1a 3dpf	The demultiplexing barcoded processing gene counting and aggregation were made using the 10x genomics' cellranger software version 6.1.2 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/output/metrics The fastq files were aligned to zebrafish genome GRCz11 and ensemble transcripts from Ensembl Release 104 by using STAR. The BAM files were as input for Cell Ranger 10X genmoics to generate processed data files that include gene names row names and cell names column names and counts. Further analysis done by using Seurat package in R. Assembly: GRCz11 Supplementary files format and content: filtered bc matrix outputs from Cell Ranger	spinal cord	No treatment done.	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics	Zebrafish were kept in the re circulating system on a 14/10 h light/dark cycle pH 7.5 at 28 °C ±1 °C in groups of 20 animals per 2.8 L.	tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:3dpf	GSM7077909	GSM7077909: sox1a 3dpf; Danio rerio; RNA Seq	GSM7077909 r1	GSM7077909	1	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP425371		assembly:GRCz11\|intentional duplicate\|dangling references:treat as unmapped	3dpf_possorted_genome_bam.bam	10X Genomics bam file	6388654202.0	63254002.0	GSM7077909 r1	0:101	A:1852098464;C:1353183909;G:1514805659;T:1668367060;N:199110	101			1852098464	1353183909	1514805659	1668367060	199110	SRX19552675	SRS16937433	SRA1601794	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	1	0.90517		0.25848		0.77025		0.56264		101		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	Germany	2023-03-02	Larval	Larval	Spinal Cord	Nervous System
74359	74359	SRR23690177	SRX19552674	SRS16937432	SRP425371	PRJNA940435	sox1a:eGFP transgenic line and single cell transcriptomics reveal the origin of zebrafish intraspinal serotonergic neurons	GSE226494	Transcriptome Analysis	The Sox family of transcription factors plays a crucial role in the development of the vertebrate nervous system. In the zebrafish embryo sox1 genes are expressed in neural progenitor cells and neurons of the ventral spinal cord. We recently reported that the loss of function of sox1a and sox1b results in a significant decrease in a subtype of V2 neurons called V2s in zebrafish. Here a single cell RNA sequencing scRNA Seq approach was used to analyse the transcriptome of sox1a lineage progenitors and neurons in the zebrafish spinal cord at four different time points during the development of an earlier established Tgsox1a:EGFP line. In addition to the sox1a expressing neurons described previously we found that this gene is also expressed in late developing intraspinal serotonin neurons ISNs. Developmental trajectory analysis of single cell data and morpholino knockdown of lateral floor plate LFP cells expressing the nkx2.9 gene suggest that ISNs are generated from LFP progenitor cells. Pharmacological inhibition of the Notch signalling pathway demonstrates the necessity of this pathway for the development of LFP progenitor cells into ISN populations. Our results confirm that the zebrafish LFP is a progenitor domain from which ISN neurons are generated in addition to the previously described KA" and V3. Overall design: The trunk and tail of Tgsox1a:eGFP lines were and dissected at 1 2 dpf 3 dpf and 5 dpf. The tissue was dissociated and GFP positive cells were sorted by FACS as described Cosacak et al. 2019. The single cell encapsulation cDNA synthesis by 10X Genomics. The reads were aligned to zebrafish genom GRChZ 11 and ensemble version 105 was used for gene annotation and assigning reads to genes. Genomics Kit and Sequencing done by Illumina.		pubmed:37529101		sox1a 2dpf	GSM7077908		source name:spinal cord\|tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:2dpf\|geo loc name:missing\|collection date:missing	sox1a 2dpf	The demultiplexing barcoded processing gene counting and aggregation were made using the 10x genomics' cellranger software version 6.1.2 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/output/metrics The fastq files were aligned to zebrafish genome GRCz11 and ensemble transcripts from Ensembl Release 104 by using STAR. The BAM files were as input for Cell Ranger 10X genmoics to generate processed data files that include gene names row names and cell names column names and counts. Further analysis done by using Seurat package in R. Assembly: GRCz11 Supplementary files format and content: filtered bc matrix outputs from Cell Ranger	spinal cord	No treatment done.	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics	Zebrafish were kept in the re circulating system on a 14/10 h light/dark cycle pH 7.5 at 28 °C ±1 °C in groups of 20 animals per 2.8 L.	tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:2dpf	GSM7077908	GSM7077908: sox1a 2dpf; Danio rerio; RNA Seq	GSM7077908 r1	GSM7077908	1	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP425371		assembly:GRCz11\|intentional duplicate\|dangling references:treat as unmapped	2dpf_possorted_genome_bam.bam	10X Genomics bam file	6112999144.0	60524744.0	GSM7077908 r1	0:101	A:1707014148;C:1361654515;G:1549767034;T:1494373967;N:189480	101			1707014148	1361654515	1549767034	1494373967	189480	SRX19552674	SRS16937432	SRA1601794	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	1	0.90304		0.26514		0.79117		0.59637		101		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	Germany	2023-03-02	Hatching	Embryo	Spinal Cord	Nervous System
74360	74360	SRR23690178	SRX19552673	SRS16937431	SRP425371	PRJNA940435	sox1a:eGFP transgenic line and single cell transcriptomics reveal the origin of zebrafish intraspinal serotonergic neurons	GSE226494	Transcriptome Analysis	The Sox family of transcription factors plays a crucial role in the development of the vertebrate nervous system. In the zebrafish embryo sox1 genes are expressed in neural progenitor cells and neurons of the ventral spinal cord. We recently reported that the loss of function of sox1a and sox1b results in a significant decrease in a subtype of V2 neurons called V2s in zebrafish. Here a single cell RNA sequencing scRNA Seq approach was used to analyse the transcriptome of sox1a lineage progenitors and neurons in the zebrafish spinal cord at four different time points during the development of an earlier established Tgsox1a:EGFP line. In addition to the sox1a expressing neurons described previously we found that this gene is also expressed in late developing intraspinal serotonin neurons ISNs. Developmental trajectory analysis of single cell data and morpholino knockdown of lateral floor plate LFP cells expressing the nkx2.9 gene suggest that ISNs are generated from LFP progenitor cells. Pharmacological inhibition of the Notch signalling pathway demonstrates the necessity of this pathway for the development of LFP progenitor cells into ISN populations. Our results confirm that the zebrafish LFP is a progenitor domain from which ISN neurons are generated in addition to the previously described KA" and V3. Overall design: The trunk and tail of Tgsox1a:eGFP lines were and dissected at 1 2 dpf 3 dpf and 5 dpf. The tissue was dissociated and GFP positive cells were sorted by FACS as described Cosacak et al. 2019. The single cell encapsulation cDNA synthesis by 10X Genomics. The reads were aligned to zebrafish genom GRChZ 11 and ensemble version 105 was used for gene annotation and assigning reads to genes. Genomics Kit and Sequencing done by Illumina.		pubmed:37529101		sox1a 1dpf	GSM7077907		source name:spinal cord\|tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:1dpf\|geo loc name:missing\|collection date:missing	sox1a 1dpf	The demultiplexing barcoded processing gene counting and aggregation were made using the 10x genomics' cellranger software version 6.1.2 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/output/metrics The fastq files were aligned to zebrafish genome GRCz11 and ensemble transcripts from Ensembl Release 104 by using STAR. The BAM files were as input for Cell Ranger 10X genmoics to generate processed data files that include gene names row names and cell names column names and counts. Further analysis done by using Seurat package in R. Assembly: GRCz11 Supplementary files format and content: filtered bc matrix outputs from Cell Ranger	spinal cord	No treatment done.	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics	Zebrafish were kept in the re circulating system on a 14/10 h light/dark cycle pH 7.5 at 28 °C ±1 °C in groups of 20 animals per 2.8 L.	tissue:spinal cord\|genotype:Tgsox1a:eGFP\|time point:1dpf	GSM7077907	GSM7077907: sox1a 1dpf; Danio rerio; RNA Seq	GSM7077907 r1	GSM7077907	1	Cells from zebrafish spinla cord were dissociated and sorted by FACS based on the reporter line. The library preparation was performed by 10X Genomics as per manufacture's protocol. 10X genomics		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500	SRP425371		intentional duplicate	1dpf_possorted_genome_bam.bam	10X Genomics bam file	41744068408.0	413307608.0	GSM7077907 r1	0:101	A:11577495181;C:9696023908;G:10077115759;T:10392411778;N:1021782	101			11577495181	9696023908	10077115759	10392411778	1021782	SRX19552673	SRS16937431	SRA1601794	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	Rastegar, Institute of Biological and Chemical Systems-Biological Information Processing (IBCS-BIP), Karlsruhe Institute of Technology (KIT)	1	0.93863		0.11011		0.8046		0.47593		101		B		usable mapping rate	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	Germany	2023-03-02	Pharyngula	Embryo	Spinal Cord	Nervous System
76472	76472	SRR24974217	SRX20731837	SRS18022738	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 6 wpi rep2	GSM7501739		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 6 wpi rep2	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501739	GSM7501739: EKAB Spinal Cord 6 wpi rep2; Danio rerio; RNA Seq	GSM7501739 r1	GSM7501739	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-EKAB_6wpi-EKAB_6wpi_060222-lib1_S1_L004_R2_001.fastq.gz MLAG-EKAB_6wpi-EKAB_6wpi_060222-lib1_S1_L004_R1_001.fastq.gz	fastq fastq	105391161150.0	592085175.0	GSM7501739 r1	0:28 1:150	A:35062246811;C:19059207172;G:22616069136;T:28650968975;N:2669056	28	150		35062246811	19059207172	22616069136	28650968975	2669056	SRX20731837	SRS18022738	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.00879	0.72202	0.00538	0.51574	0.99452	0.83798	0.28129	0.53924	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76473	76473	SRR24974218	SRX20731836	SRS18022737	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 6 wpi rep1	GSM7501738		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 6 wpi rep1	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501738	GSM7501738: EKAB Spinal Cord 6 wpi rep1; Danio rerio; RNA Seq	GSM7501738 r1	GSM7501738	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-6wpi-EKAB-6wpi_EKAB_061621-lib1_S1_L003_R2_001.fastq.gz MLAG-6wpi-EKAB-6wpi_EKAB_061621-lib1_S1_L003_R1_001.fastq.gz	fastq fastq	84895623668.0	476941706.0	GSM7501738 r1	0:28 1:150	A:26734879880;C:16050029642;G:18461921593;T:23647200445;N:1592108	28	150		26734879880	16050029642	18461921593	23647200445	1592108	SRX20731836	SRS18022737	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.01259	0.78757	0.00729	0.49153	0.99511	0.83721	0.27956	0.64396	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76474	76474	SRR24974219	SRX20731835	SRS18022736	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 3 wpi rep2	GSM7501737		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 3 wpi rep2	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501737	GSM7501737: EKAB Spinal Cord 3 wpi rep2; Danio rerio; RNA Seq	GSM7501737 r1	GSM7501737	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-EKAB_3wpi-EKAB_3wpi_060222-lib1_S2_L004_R2_001.fastq.gz MLAG-EKAB_3wpi-EKAB_3wpi_060222-lib1_S2_L004_R1_001.fastq.gz	fastq fastq	101322905824.0	569229808.0	GSM7501737 r1	0:28 1:150	A:33529740337;C:18508795541;G:22218128467;T:27063600389;N:2641090	28	150		33529740337	18508795541	22218128467	27063600389	2641090	SRX20731835	SRS18022736	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.01016	0.72116	0.00605	0.48692	0.99462	0.83666	0.28172	0.53814	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76475	76475	SRR24974220	SRX20731834	SRS18022735	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 3 wpi rep1	GSM7501736		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 3 wpi rep1	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501736	GSM7501736: EKAB Spinal Cord 3 wpi rep1; Danio rerio; RNA Seq	GSM7501736 r1	GSM7501736	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-3wpi-EKAB-3wpi_EKAB_081121-lib1_S1_L003_R1_001.fastq.gz MLAG-3wpi-EKAB-3wpi_EKAB_081121-lib1_S1_L003_R2_001.fastq.gz	fastq fastq	105936769400.0	529683847.0	GSM7501736 r1	0:50 1:150	A:35005213053;C:19468074725;G:21726475457;T:29732821194;N:4184971	50	150		35005213053	19468074725	21726475457	29732821194	4184971	SRX20731834	SRS18022735	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.0	0.74755	0.0	0.45633	1.0	0.81667		0.52945	50	150	T	B	mate1 technical by mapping diff	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76476	76476	SRR24974221	SRX20731833	SRS18022734	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 1 wpi rep2	GSM7501735		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 1 wpi rep2	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501735	GSM7501735: EKAB Spinal Cord 1 wpi rep2; Danio rerio; RNA Seq	GSM7501735 r1	GSM7501735	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-EKAB_1wpi-EKAB_1wpi_060122-lib1_S4_L004_R1_001.fastq.gz MLAG-EKAB_1wpi-EKAB_1wpi_060122-lib1_S4_L004_R2_001.fastq.gz	fastq fastq	74288465184.0	417350928.0	GSM7501735 r1	0:28 1:150	A:23894773694;C:13783822381;G:16246726428;T:20361238077;N:1904604	28	150		23894773694	13783822381	16246726428	20361238077	1904604	SRX20731833	SRS18022734	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.01015	0.74263	0.00625	0.51946	0.99464	0.84062	0.29395	0.55744	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76477	76477	SRR24974222	SRX20731832	SRS18022733	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord 1 wpi rep1	GSM7501734		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord 1 wpi rep1	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Injured	GSM7501734	GSM7501734: EKAB Spinal Cord 1 wpi rep1; Danio rerio; RNA Seq	GSM7501734 r1	GSM7501734	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-1wpi-EKAB-1wpi_EKAB_022521_S1_L001_R1_001.fastq.gz MLAG-1wpi-EKAB-1wpi_EKAB_022521_S1_L001_R2_001.fastq.gz	fastq fastq	105664571464.0	593621188.0	GSM7501734 r1	0:28 1:150	A:36626846226;C:18608728346;G:22744330287;T:27679941436;N:4725169	28	150		36626846226	18608728346	22744330287	27679941436	4725169	SRX20731832	SRS18022733	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.01645	0.6943	0.01249	0.36909	0.9919	0.8309	0.33377	0.52756	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76478	76478	SRR24974223	SRX20731831	SRS18022732	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord Uninjured rep2	GSM7501733		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Uninjured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord Uninjured rep2	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Uninjured	GSM7501733	GSM7501733: EKAB Spinal Cord Uninjured rep2; Danio rerio; RNA Seq	GSM7501733 r1	GSM7501733	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-EKAB_Uninj-EKAB_Uninj_060122-lib1_S3_L003_R1_001.fastq.gz MLAG-EKAB_Uninj-EKAB_Uninj_060122-lib1_S3_L003_R2_001.fastq.gz	fastq fastq	70360166986.0	395281837.0	GSM7501733 r1	0:28 1:150	A:23061071505;C:12770839971;G:14766106153;T:19760221018;N:1928339	28	150		23061071505	12770839971	14766106153	19760221018	1928339	SRX20731831	SRS18022732	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.00827	0.72918	0.00547	0.5217	0.99415	0.82402	0.31226	0.5218	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System
76479	76479	SRR24974224	SRX20731830	SRS18022731	SRP444940	PRJNA985790	Single cell analysis of innate spinal cord regeneration identifies intersecting modes of neuronal repair	GSE235395	Transcriptome Analysis	Adult zebrafish have an innate ability to recover from severe spinal cord injury. Here we report a comprehensive single nuclear RNA sequencing atlas that spans 6 weeks of regeneration. We identify cooperative roles for adult neurogenesis and neuronal plasticity during spinal cord repair. Neurogenesis of glutamatergic and GABAergic neurons restores the excitatory/inhibitory xxx post injury. In addition a transient population of injury responsive neurons iNeurons show elevated plasticity between 1 and 3 xxx post injury. We found iNeurons are injury surviving neurons that acquire a neuroblast like gene expression xxx post injury. CRISPR/Cas9 mutagenesis showed iNeurons are required for functional recovery and employ vesicular trafficking as an essential mechanism that underlies neuronal plasticity. This study provides a comprehensive resource of the cells and mechanisms that direct spinal cord regeneration and establishes zebrafish as a model of plasticity driven neural repair. Overall design: we performed complete SC transections on adult zebrafish and dissected 3 mm sections of SC tissues surrounding the lesion site at 1 3 and 6 xxx post injury wpi for nuclear isolation. Corresponding tissue sections were collected from uninjured controls. SC tissues were pooled from 50 animals per time point and 2 pools of independent biological replicates were analyzed for each time point. Our dataset spans key regenerative windows including early injury induced signals at 1 wpi neuronal and glial regeneration at 3 wpi and cellular remodeling at late stages of regeneration at 6 wpi Mokalled et al. 2016. Isolated nuclei were sequenced using 10x genomics platform three prime v3.1 chemistry and aligned to zebrafish genome GRCz11 with improved zebrafish transcriptome annotation Lawson et al. 2020; Matson et al. 2018. Nuclei were subsequently filtered using the Decontx and DoubletFinder packages to exclude droplets that include doublet nuclei or a high proportion of ambient mRNA respectively McGinnis et al. 2019; Yang et …		pubmed:39147780		EKAB Spinal Cord Uninjured rep1	GSM7501732		source name:Spinal Cord\|tissue:Spinal Cord\|genotype:Wild type\|treatment:Uninjured\|geo loc name:missing\|collection date:missing	EKAB Spinal Cord Uninjured rep1	post sequencing the Illumina output was processed using the CellRanger v6.0.0 recommended pipeline to generate gene barcode count matrices. A custom reference genome was made with the “cellranger mkref” command using the fasta file of zebrafish reference genome GRCz11 constructed from the Ensemble genome build https://useast.ensembl.org/Danio rerio/Info/Index and the sorted Gene Transfer Format file v4.3.2 from the improved zebrafish transcriptome annotation Lawson et al. 2020 Base call files for each sample from Illumina were demultiplexed into FASTQ reads. Then the “cellranger count” pipeline was used to align sequencing reads in FASTQ files to the custom reference genome. Both exon and intron sequences were included during the alignment. The filtered gene barcode count matrices generated by “cellranger count” was used for downstream analysis. All the datasets were integrated and analyzed using Seurat v4.1.1 package with R v4.2.1 R Core Team 2018; Stuart et al. 2019. Each sample count matrix was filtered for genes that were expressed in at least 3 cells and cells expressing at least 200 genes followed by cell quality assessment using commonly used QC matrixes Ilicic et al. 2016. Cells having a unique number of genes between 200 to 4000 and a mitochondrial gene percentage <5 were used for downstream processing. Each dataset was independently normalized and scaled using the “SCTransform” function which is an improved method for normalization that performs a variance stabilizing transformation using negative binomial regression Hafemeister and Satija 2019. Standard integration workflow of Seurat was used to identify shared sources of variation across experiments as well as mutual nearest neighbors Butler et al. 2018; Haghverdi et al. 2018. Integration features were selected based on the top 4000 highly variable features using “SelectIntegrationFeatures” function nfeatures = 4000 which was used as input for the “anchor.features” argument of the “FindIntegrationAnchors” function. PCA analysis was per…	Spinal Cord		For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell 3’ Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell 3’ Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer’s protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		tissue:Spinal Cord\|genotype:Wild type\|treatment:Uninjured	GSM7501732	GSM7501732: EKAB Spinal Cord Uninjured rep1; Danio rerio; RNA Seq	GSM7501732 r1	GSM7501732	1	For snRNA seq 30 µl of resuspension solution containing isolated nuclei at a concentration of 1000 nuclei/µl was submitted to sequencing.Two biological replicates of each timepoints at 0 1 3 and 6 wpi were used. cDNA was prepared post the GEM generation and barcoding followed by the GEM RT reaction and bead cleanup steps. cDNA was amplified for 11 13 cycles then purified using SPRIselect beads. Purified cDNA samples were then run on a Bioanalyzer to determine the cDNA concentration. GEX libraries were prepared as recommended by the 10x Genomics Chromium Single Cell three prime Reagent Kits User Guide v3.1 Chemistry Dual Index with appropriate modifications to the PCR cycles based on the calculated cDNA concentration. For sample preparation on the 10x Genomics platform the Chromium Next GEM Single Cell three prime Kit v3.1 16 rxns PN 1000268 Chromium Next GEM Chip G Single Cell Kit 48 rxns PN 1000120 and Dual Index Kit TT Set A 96 rxns PN 1000215 were used. The concentration of each library was accurately determined through qPCR utilizing the KAPA library Quantification Kit according to the manufacturer's protocol KAPA Biosystems/Roche to produce cluster counts appropriate for the Illumina NovaSeq6000 instrument. Normalized libraries were sequenced on a NovaSeq6000 S4 Flow Cell using the XP workflow and a 50x10x16x150 sequencing recipe according to manufacturer protocol. A median sequencing depth of 50 000 reads/cell was targeted for each Gene Expression Library.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000	SRP444940		loader:fastq load.py	MLAG-Uninj-EKAB-Uninj_EKAB_022521_S2_L001_R1_001.fastq.gz MLAG-Uninj-EKAB-Uninj_EKAB_022521_S2_L001_R2_001.fastq.gz	fastq fastq	90709334000.0	509603000.0	GSM7501732 r1	0:28 1:150	A:31112543559;C:15894186068;G:19250857481;T:24447634392;N:4112500	28	150		31112543559	15894186068	19250857481	24447634392	4112500	SRX20731830	SRS18022731	SRA1659265	Mokalled Lab, Developmental Biology, Washington University in St. Louis	Developmental Biology, Washington University in St. Louis	2	0.01435	0.67925	0.01151	0.41035	0.99397	0.83936	0.36598	0.58394	28	150	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x	United States	2023-06-20	Undetermined	Adult	Spinal Cord	Nervous System

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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");;