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.spot_descriptor,experiment.study_ref,run.title,run.attributes,run.filename,run.semantic_name,run.total_bases,run.total_spots,run.alias,run.read_lengths,run.base_counts,run.r1_length,run.r2_length,run.r3_length,run.r4_length,run.Acount,run.Ccount,run.Gcount,run.Tcount,run.Ncount,run.experiment,run.pool_member,submission.accession,submission.srasource,submission.bioprojectsource,seqdetective.n_mates,seqdetective.mapping_rate.mate1,seqdetective.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,tech_variant,submission.bioprojectsource.country,earliest_date,devstage_curation,devstage_curation_coarse,tissue_curation,tissue_curation_coarse
15009,ERR12306826,ERX11683773,ERS17043893,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Coated,SAMEA114641726,KIS,ENA first public:2024 09 30|External Id:SAMEA114641726|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Coated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Coated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Coated p,Coated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Coated_S1_L003_I1_001.fastq.gz 230324-Coated_S1_L003_I2_001.fastq.gz 230324-Coated_S1_L003_R1_001.fastq.gz 230324-Coated_S1_L003_R2_001.fastq.gz,fastq fastq fastq fastq,10050750400.0,62817190.0,E MTAB 13554:230324 Coated S1 L003,0:10 1:10 2:30 3:110,A:1894211982;C:1556769050;G:1595134885;T:1863244619;N:530364,10,10,30,110,1894211982,1556769050,1595134885,1863244619,530364,ERX11683773,ERS17043893,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15010,ERR12306829,ERX11683773,ERS17043893,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Coated,SAMEA114641726,KIS,ENA first public:2024 09 30|External Id:SAMEA114641726|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Coated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Coated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Coated p,Coated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Coated_S1_L004_I1_001.fastq.gz 230324-Coated_S1_L004_I2_001.fastq.gz 230324-Coated_S1_L004_R1_001.fastq.gz 230324-Coated_S1_L004_R2_001.fastq.gz,fastq fastq fastq fastq,9780757760.0,61129736.0,E MTAB 13554:230324 Coated S1 L004,0:10 1:10 2:30 3:110,A:1844808667;C:1516149264;G:1549838165;T:1813091860;N:383004,10,10,30,110,1844808667,1516149264,1549838165,1813091860,383004,ERX11683773,ERS17043893,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15011,ERR12306828,ERX11683773,ERS17043893,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Coated,SAMEA114641726,KIS,ENA first public:2024 09 30|External Id:SAMEA114641726|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Coated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Coated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Coated p,Coated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Coated_S1_L002_I1_001.fastq.gz 230324-Coated_S1_L002_I2_001.fastq.gz 230324-Coated_S1_L002_R1_001.fastq.gz 230324-Coated_S1_L002_R2_001.fastq.gz,fastq fastq fastq fastq,9639368480.0,60246053.0,E MTAB 13554:230324 Coated S1 L002,0:10 1:10 2:30 3:110,A:1817388578;C:1494319441;G:1527564319;T:1787447672;N:345820,10,10,30,110,1817388578,1494319441,1527564319,1787447672,345820,ERX11683773,ERS17043893,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15012,ERR12306832,ERX11683773,ERS17043893,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Coated,SAMEA114641726,KIS,ENA first public:2024 09 30|External Id:SAMEA114641726|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Coated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Coated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Coated p,Coated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Coated_S1_L001_I1_001.fastq.gz 230324-Coated_S1_L001_I2_001.fastq.gz 230324-Coated_S1_L001_R1_001.fastq.gz 230324-Coated_S1_L001_R2_001.fastq.gz,fastq fastq fastq fastq,9823147840.0,61394674.0,E MTAB 13554:230324 Coated S1 L001,0:10 1:10 2:30 3:110,A:1851113558;C:1521671335;G:1558230628;T:1822067428;N:331191,10,10,30,110,1851113558,1521671335,1558230628,1822067428,331191,ERX11683773,ERS17043893,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15013,ERR12306833,ERX11683774,ERS17043894,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Uncoated,SAMEA114641727,KIS,ENA first public:2024 09 30|External Id:SAMEA114641727|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Uncoated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Uncoated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Uncoated p,Uncoated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Uncoated_S2_L001_I1_001.fastq.gz 230324-Uncoated_S2_L001_I2_001.fastq.gz 230324-Uncoated_S2_L001_R1_001.fastq.gz 230324-Uncoated_S2_L001_R2_001.fastq.gz,fastq fastq fastq fastq,8871716800.0,55448230.0,E MTAB 13554:230324 Uncoated S2 L001,0:10 1:10 2:30 3:110,A:1718985845;C:1319560520;G:1372197713;T:1688254655;N:306567,10,10,30,110,1718985845,1319560520,1372197713,1688254655,306567,ERX11683774,ERS17043894,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15014,ERR12306830,ERX11683774,ERS17043894,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Uncoated,SAMEA114641727,KIS,ENA first public:2024 09 30|External Id:SAMEA114641727|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Uncoated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Uncoated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Uncoated p,Uncoated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Uncoated_S2_L003_I1_001.fastq.gz 230324-Uncoated_S2_L003_I2_001.fastq.gz 230324-Uncoated_S2_L003_R1_001.fastq.gz 230324-Uncoated_S2_L003_R2_001.fastq.gz,fastq fastq fastq fastq,9075478720.0,56721742.0,E MTAB 13554:230324 Uncoated S2 L003,0:10 1:10 2:30 3:110,A:1758677834;C:1349774831;G:1404463433;T:1725994897;N:480625,10,10,30,110,1758677834,1349774831,1404463433,1725994897,480625,ERX11683774,ERS17043894,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15015,ERR12306831,ERX11683774,ERS17043894,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Uncoated,SAMEA114641727,KIS,ENA first public:2024 09 30|External Id:SAMEA114641727|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Uncoated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Uncoated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Uncoated p,Uncoated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Uncoated_S2_L002_I1_001.fastq.gz 230324-Uncoated_S2_L002_I2_001.fastq.gz 230324-Uncoated_S2_L002_R1_001.fastq.gz 230324-Uncoated_S2_L002_R2_001.fastq.gz,fastq fastq fastq fastq,8705210880.0,54407568.0,E MTAB 13554:230324 Uncoated S2 L002,0:10 1:10 2:30 3:110,A:1687330583;C:1295837902;G:1345522657;T:1655826422;N:314916,10,10,30,110,1687330583,1295837902,1345522657,1655826422,314916,ERX11683774,ERS17043894,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
15016,ERR12306827,ERX11683774,ERS17043894,ERP155237,PRJEB70303,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E-MTAB-13554,Transcriptome Analysis,To study the biodistribution of DNA origami in zebrafish and to determine differences in biodistribution based on the coating of the nanostructure using oligolysine PEG copolymers  zebrafish embryos 2dpf were injected with coated or uncoated DNA origami wireframe nanosheets without xxx targeting molecule  but containing a detection fluorophores. post treatment  the embryos were disassociated into a single cell suspension and sorted through FACS for embryos containing nanostructures fluorophores. 10X sc libraries were generated from the sorted embryos and sequenced to establish cell type predictions for cells with coated and uncoated structures respectively.,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,,Protocols: Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences. Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd. Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells. The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,Uncoated,SAMEA114641727,KIS,ENA first public:2024 09 30|External Id:SAMEA114641727|INSDC center name:KIS|INSDC last update:2023 11 20T16:06:50Z|INSDC status:public|Submitter Id:E MTAB 13554:Uncoated|age:2|broker name:ArrayExpress|collection date:not collected|common name:zebrafish|developmental stage:embryo stage|geographic location country and/or sea:not collected|isolate:not applicable|organism part:whole organism|sample name:E MTAB 13554:Uncoated|scientific name:Danio rerio|sex:not available|strain:transgenic lines Tgfli1:EGFPy1,,,,,,,,,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,E MTAB 13554:Uncoated p,Uncoated p,10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,Embryos 48 hpf were disassociated into a single cell suspension. Four hours post injections  around 200 Tgfli1:EGFP embryos were euthanized in 0.02% tricaine. The tissue was dissociated with harsh pipetting in 0.25% trypsin EDTA GIBCO and 100mg/mL collagenase Sigma Aldrich for 3 min at 30oC  followed by resuspension in DMEM 10% FBS. The cells were then pelleted at 700xg for 5 min  washed once in 1X PBS  filtered through a 40 μm cell strainer MERCK and stained with 1mM DAPI solution Abcam. TS+ cells were FACS sorted on a FACSARIA III with BD FACSDIVA software v. 9.0.1 BD Biosciences.  Zebrafish D. rerio were maintained and bred in compliance with Swedish legislation on animal welfare regulations approved by Stockholms djurförsöksetiska nämnd.  Each zebrafish embryo Tgfli1:EGFP was microinjected with 4 nl of the sample. The samples for injections were prepared at a final concentration of 100 nM in 1x PBS effective dose estimated to around 4 6 nM. Embryos at 48 hpf were anesthetized using 0.1% tricaine MS222  transferred onto an agarose surface  placed on their ventral side  and injected into the common cardinal vein Duct of Cuvier. scRNAseq libraries were generated from the sorted cells using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 per manufacturer's instructions  targeting 8000 cells.  The cDNA libraries were amplified using 10X Genomics Chromium Single Cell 3ʹ Reagent Kits v3.1 with 14 PCR cycles.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,PolyA,PAIRED,ILLUMINA,NextSeq 550,,ERP155237,NextSeq 550 paired end sequencing; 10X genomics libraries for scRNA seq of zebrafish embryos 2dpf treated with uncoated and oligolysine PEG copolymer coated DNA origami wireframe nanosheets,ENA FIRST PUBLIC:2024 09 30|ENA LAST UPDATE:2024 09 30,230324-Uncoated_S2_L004_I1_001.fastq.gz 230324-Uncoated_S2_L004_I2_001.fastq.gz 230324-Uncoated_S2_L004_R1_001.fastq.gz 230324-Uncoated_S2_L004_R2_001.fastq.gz,fastq fastq fastq fastq,8846203360.0,55288771.0,E MTAB 13554:230324 Uncoated S2 L004,0:10 1:10 2:30 3:110,A:1715100047;C:1316853239;G:1367208350;T:1682248092;N:355082,10,10,30,110,1715100047,1316853239,1367208350,1682248092,355082,ERX11683774,ERS17043894,ERA27417303,European Bioinformatics Institute|European Nucleotide Archive,European Bioinformatics Institute|European Nucleotide Archive,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq,unknown,poly_a,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-11-20,Hatching,Embryo,Whole Organism,All anatomical structures
24838,SRR25509990,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S65_L001_I1_001.fastq.gz Kristen_10X_101620_S65_L001_R1_001.fastq.gz Kristen_10X_101620_S65_L001_R2_001.fastq.gz,fastq fastq fastq,3327423622.0,26200186.0,GSM7680082 r1,0:8 1:28 2:91,A:693698335;C:494975421;G:540414923;T:654926705;N:201542,8,28,91,,693698335,494975421,540414923,654926705,201542,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91351,,0.22865,,0.77723,,0.49805,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24839,SRR25509991,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S67_L002_I1_001.fastq.gz Kristen_10X_101620_S67_L002_R1_001.fastq.gz Kristen_10X_101620_S67_L002_R2_001.fastq.gz,fastq fastq fastq,4430975390.0,34889570.0,GSM7680082 r10,0:8 1:28 2:91,A:923557751;C:658731986;G:718325003;T:874072049;N:264081,8,28,91,,923557751,658731986,718325003,874072049,264081,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91797,,0.22868,,0.77863,,0.50737,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24840,SRR25509992,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S67_L003_I1_001.fastq.gz Kristen_10X_101620_S67_L003_R1_001.fastq.gz Kristen_10X_101620_S67_L003_R2_001.fastq.gz,fastq fastq fastq,4429223552.0,34875776.0,GSM7680082 r11,0:8 1:28 2:91,A:923277621;C:658440918;G:718036099;T:873676803;N:264175,8,28,91,,923277621,658440918,718036099,873676803,264175,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91753,,0.22978,,0.77942,,0.50977,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24841,SRR25509993,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S67_L004_I1_001.fastq.gz Kristen_10X_101620_S67_L004_R1_001.fastq.gz Kristen_10X_101620_S67_L004_R2_001.fastq.gz,fastq fastq fastq,4549656890.0,35824070.0,GSM7680082 r12,0:8 1:28 2:91,A:950423951;C:673969790;G:734760168;T:900554096;N:282365,8,28,91,,950423951,673969790,734760168,900554096,282365,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91687,,0.23323,,0.77932,,0.50984,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24842,SRR25509994,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S68_L001_I1_001.fastq.gz Kristen_10X_101620_S68_L001_R1_001.fastq.gz Kristen_10X_101620_S68_L001_R2_001.fastq.gz,fastq fastq fastq,2696456634.0,21231942.0,GSM7680082 r13,0:8 1:28 2:91,A:562209710;C:401113540;G:437836456;T:530785441;N:161575,8,28,91,,562209710,401113540,437836456,530785441,161575,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91384,,0.22885,,0.77912,,0.50672,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24843,SRR25509995,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S68_L002_I1_001.fastq.gz Kristen_10X_101620_S68_L002_R1_001.fastq.gz Kristen_10X_101620_S68_L002_R2_001.fastq.gz,fastq fastq fastq,2735162932.0,21536716.0,GSM7680082 r14,0:8 1:28 2:91,A:570169460;C:406846839;G:443964376;T:538696359;N:164122,8,28,91,,570169460,406846839,443964376,538696359,164122,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91691,,0.22998,,0.77942,,0.51035,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24844,SRR25509996,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S68_L003_I1_001.fastq.gz Kristen_10X_101620_S68_L003_R1_001.fastq.gz Kristen_10X_101620_S68_L003_R2_001.fastq.gz,fastq fastq fastq,2739703563.0,21572469.0,GSM7680082 r15,0:8 1:28 2:91,A:571283074;C:407511119;G:444574049;T:539563747;N:162690,8,28,91,,571283074,407511119,444574049,539563747,162690,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91526,,0.22747,,0.7793,,0.51387,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24845,SRR25509997,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S68_L004_I1_001.fastq.gz Kristen_10X_101620_S68_L004_R1_001.fastq.gz Kristen_10X_101620_S68_L004_R2_001.fastq.gz,fastq fastq fastq,2828960433.0,22275279.0,GSM7680082 r16,0:8 1:28 2:91,A:591166850;C:419193897;G:457332081;T:559181603;N:175958,8,28,91,,591166850,419193897,457332081,559181603,175958,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91651,,0.23126,,0.77613,,0.50223,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24846,SRR25509998,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S65_L002_I1_001.fastq.gz Kristen_10X_101620_S65_L002_R1_001.fastq.gz Kristen_10X_101620_S65_L002_R2_001.fastq.gz,fastq fastq fastq,3374189340.0,26568420.0,GSM7680082 r2,0:8 1:28 2:91,A:703424426;C:501911907;G:547786373;T:664402117;N:201397,8,28,91,,703424426,501911907,547786373,664402117,201397,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91522,,0.22812,,0.77753,,0.50208,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24847,SRR25509999,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S65_L003_I1_001.fastq.gz Kristen_10X_101620_S65_L003_R1_001.fastq.gz Kristen_10X_101620_S65_L003_R2_001.fastq.gz,fastq fastq fastq,3383217516.0,26639508.0,GSM7680082 r3,0:8 1:28 2:91,A:705514047;C:503189161;G:549089218;T:666200894;N:201908,8,28,91,,705514047,503189161,549089218,666200894,201908,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91535,,0.23036,,0.78078,,0.50202,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24848,SRR25510000,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S65_L004_I1_001.fastq.gz Kristen_10X_101620_S65_L004_R1_001.fastq.gz Kristen_10X_101620_S65_L004_R2_001.fastq.gz,fastq fastq fastq,3493154050.0,27505150.0,GSM7680082 r4,0:8 1:28 2:91,A:729910298;C:517582902;G:564926384;T:690333365;N:215701,8,28,91,,729910298,517582902,564926384,690333365,215701,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91569,,0.22913,,0.77873,,0.50186,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24849,SRR25510001,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S66_L001_I1_001.fastq.gz Kristen_10X_101620_S66_L001_R1_001.fastq.gz Kristen_10X_101620_S66_L001_R2_001.fastq.gz,fastq fastq fastq,3237456187.0,25491781.0,GSM7680082 r5,0:8 1:28 2:91,A:674885196;C:481871349;G:525642933;T:637154812;N:197781,8,28,91,,674885196,481871349,525642933,637154812,197781,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91378,,0.22852,,0.77843,,0.50985,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24850,SRR25510002,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S66_L002_I1_001.fastq.gz Kristen_10X_101620_S66_L002_R1_001.fastq.gz Kristen_10X_101620_S66_L002_R2_001.fastq.gz,fastq fastq fastq,3287789081.0,25888103.0,GSM7680082 r6,0:8 1:28 2:91,A:685408765;C:489232060;G:533550966;T:647427897;N:197685,8,28,91,,685408765,489232060,533550966,647427897,197685,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91484,,0.22872,,0.78001,,0.49909,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24851,SRR25510003,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S66_L003_I1_001.fastq.gz Kristen_10X_101620_S66_L003_R1_001.fastq.gz Kristen_10X_101620_S66_L003_R2_001.fastq.gz,fastq fastq fastq,3298207145.0,25970135.0,GSM7680082 r7,0:8 1:28 2:91,A:687621787;C:490851045;G:535230137;T:649382247;N:197069,8,28,91,,687621787,490851045,535230137,649382247,197069,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91559,,0.2268,,0.77766,,0.50935,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24852,SRR25510004,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S66_L004_I1_001.fastq.gz Kristen_10X_101620_S66_L004_R1_001.fastq.gz Kristen_10X_101620_S66_L004_R2_001.fastq.gz,fastq fastq fastq,3377888088.0,26597544.0,GSM7680082 r8,0:8 1:28 2:91,A:705624374;C:500985504;G:546306692;T:667248332;N:211602,8,28,91,,705624374,500985504,546306692,667248332,211602,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91679,,0.22965,,0.77837,,0.51495,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
24853,SRR25510005,SRX21240518,SRS18495441,SRP453227,PRJNA1001940,Determinants of Motor Neuron Functional Subtypes Important for Locomotor Speed,GSE240026,Transcriptome Analysis,Locomotion requires precise control of the strength and speed of muscle contraction and is achieved by recruiting functionally distinct subtypes of motor neurons MNs. MNs are essential to movement and differentially susceptible in disease  but little is known about how MNs acquire functional subtype specific features during development. Using single cell RNA profiling in embryonic and larval zebrafish  we identify novel and conserved molecular signatures for MN functional subtypes  and identify genes expressed in both early post mitotic and mature MNs. Assessing MN development in genetic mutants  we define a molecular program essential for MN functional subtype specification. Two evolutionarily conserved transcription factors  Prdm16 and Mecom  are both functional subtype specific determinants integral for fast MN development. Loss of prdm16 or mecom causes fast MNs to develop transcriptional profiles and innervation similar to slow MNs. These results reveal the molecular diversity of vertebrate axial MNs and demonstrate that functional subtypes are specified through intrinsic transcriptional codes. Overall design: Spinal cord MNs of WT  prdm16 /   and mecom  /  zebrafish embryos at two developmental timepoints 2dpf and 5dpf were isolated by fluorescence activated cell sorting FACS according to the presence of dsRed and GFP signal  then analyzed using scRNAseq.,,pubmed:37676768,,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,GSM7680082,,source name:tail|tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs|geo loc name:missing|collection date:missing,Spinal cord MNs  olig2dsRed  2dpf  scRNAseq,The demultiplexing  barcoded processing  gene counting and aggregation were made using the Cell Ranger software v5.0.1 https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger Assembly: build GRCz11 Supplementary files format and content: h5 files,tail,,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter’s instructions single cell 3’ v2 protocol  10x Genomics.,,tissue:tail|genotype:olig2:dsRed  prdm16gal4:UASGFP|age:2dpf|cell type:Spinal cord MNs,GSM7680082,GSM7680082: Spinal cord MNs  olig2dsRed  2dpf  scRNAseq; Danio rerio; RNA Seq,GSM7680082 r1,GSM7680082,1,DsRed positive cells were isolated from 2 dpf or 5 dpf control  mecom mutant  or prdm16 mutant zebrafish embryos on an Tgolig2:DsRed2 or Tgprdm16::GFP;Tgolig2:DsRed2 background. Trunk and tail tissue were separated from cranial tissue. Tissue was then finely chopped using a razor blade  dissociated using papain  filtered  and resuspended for sorting. Cells were sorted using the Sony SH800 FACS Cell Sorter. GFP   DsRed2   and single fluorophore control embryos were also included as controls for the FACS setup. DAPI was used for a live/dead marker. Sorted cells were then counted using a hemocytometer and spun down to resuspend at a higher concentration. Cells were then processed using the standard 10x Genomics and CellRangerv5.0.1 pipeline 89. Raw sequencing reads were mapped to the zebrafish reference genome build GRCz11. Library was performed according to the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP453227,,loader:fastq load.py,Kristen_10X_101620_S67_L001_I1_001.fastq.gz Kristen_10X_101620_S67_L001_R1_001.fastq.gz Kristen_10X_101620_S67_L001_R2_001.fastq.gz,fastq fastq fastq,4346114117.0,34221371.0,GSM7680082 r9,0:8 1:28 2:91,A:905838814;C:646261347;G:704993570;T:856789016;N:262014,8,28,91,,905838814,646261347,704993570,856789016,262014,SRX21240518,SRS18495441,SRA1686067,"Dasen, Neuroscience Institute, New York University Grossman School of Medicine","Dasen, Neuroscience Institute, New York University Grossman School of Medicine",1,0.91558,,0.22834,,0.7783,,0.49972,,91,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2023-08-03,Hatching,Embryo,Tail,Multi-system
25107,SRR25605432,SRX21332628,SRS18578260,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep3,GSM7702835,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702835,GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq,GSM7702835 r1,GSM7702835,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_GFP_pos_S4_L001_R2_001.fastq.gz 2707_GFP_pos_S4_L001_R1_001.fastq.gz 2707_GFP_pos_S4_L001_I2_001.fastq.gz 2707_GFP_pos_S4_L001_I1_001.fastq.gz,fastq fastq fastq fastq,21603309774.0,156545723.0,GSM7702835 r1,0:10 1:10 2:28 3:90,A:3960451458;C:3105338977;G:3480072824;T:3541218595;N:2033216,10,10,28,90,3960451458,3105338977,3480072824,3541218595,2033216,SRX21332628,SRS18578260,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94286,,0.14175,,0.78395,,0.51541,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25108,SRR25605433,SRX21332628,SRS18578260,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep3,GSM7702835,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702835,GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq,GSM7702835 r1,GSM7702835,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_GFP_pos_S4_L002_I1_001.fastq.gz 2707_GFP_pos_S4_L002_I2_001.fastq.gz 2707_GFP_pos_S4_L002_R1_001.fastq.gz 2707_GFP_pos_S4_L002_R2_001.fastq.gz,fastq fastq fastq fastq,22413465270.0,162416415.0,GSM7702835 r2,0:10 1:10 2:28 3:90,A:4104405257;C:3220235633;G:3625448968;T:3667329471;N:58021,10,10,28,90,4104405257,3220235633,3625448968,3667329471,58021,SRX21332628,SRS18578260,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94239,,0.14234,,0.78338,,0.50616,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25109,SRR25822232,SRX21332628,SRS18578260,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep3,GSM7702835,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702835,GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq,GSM7702835 r1,GSM7702835,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_GFP_pos_S4_L001_R2_002.fastq.gz 2707_GFP_pos_S4_L001_R1_002.fastq.gz 2707_GFP_pos_S4_L001_I2_002.fastq.gz 2707_GFP_pos_S4_L001_I1_002.fastq.gz,fastq fastq fastq fastq,21424338264.0,155248828.0,GSM7702835 r3,0:10 1:10 2:28 3:90,A:3928077020;C:3077672525;G:3451667186;T:3513734857;N:1242932,10,10,28,90,3928077020,3077672525,3451667186,3513734857,1242932,SRX21332628,SRS18578260,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.9433,,0.14242,,0.78328,,0.52384,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25110,SRR25822233,SRX21332628,SRS18578260,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep3,GSM7702835,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702835,GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq,GSM7702835 r1,GSM7702835,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_GFP_pos_S4_L002_I1_002.fastq.gz 2707_GFP_pos_S4_L002_I2_002.fastq.gz 2707_GFP_pos_S4_L002_R1_002.fastq.gz 2707_GFP_pos_S4_L002_R2_002.fastq.gz,fastq fastq fastq fastq,22542479952.0,163351304.0,GSM7702835 r4,0:10 1:10 2:28 3:90,A:4130174287;C:3235362228;G:3643457599;T:3691295761;N:1327485,10,10,28,90,4130174287,3235362228,3643457599,3691295761,1327485,SRX21332628,SRS18578260,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94183,,0.14124,,0.78301,,0.50517,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25111,SRR25605434,SRX21332627,SRS18578259,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep3,GSM7702834,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702834,GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq,GSM7702834 r1,GSM7702834,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_pos_Mcherry_S3_L001_R2_001.fastq.gz 2707_pos_Mcherry_S3_L001_R1_001.fastq.gz 2707_pos_Mcherry_S3_L001_I2_001.fastq.gz 2707_pos_Mcherry_S3_L001_I1_001.fastq.gz,fastq fastq fastq fastq,20161522758.0,146097991.0,GSM7702834 r1,0:10 1:10 2:28 3:90,A:3649819934;C:2958286158;G:3267504300;T:3271384686;N:1824112,10,10,28,90,3649819934,2958286158,3267504300,3271384686,1824112,SRX21332627,SRS18578259,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94851,,0.1483,,0.80012,,0.52549,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25112,SRR25605435,SRX21332627,SRS18578259,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep3,GSM7702834,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702834,GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq,GSM7702834 r1,GSM7702834,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_pos_Mcherry_S3_L002_I1_001.fastq.gz 2707_pos_Mcherry_S3_L002_I2_001.fastq.gz 2707_pos_Mcherry_S3_L002_R1_001.fastq.gz 2707_pos_Mcherry_S3_L002_R2_001.fastq.gz,fastq fastq fastq fastq,20708273928.0,150059956.0,GSM7702834 r2,0:10 1:10 2:28 3:90,A:3746088654;C:3037032642;G:3368938151;T:3353289242;N:47351,10,10,28,90,3746088654,3037032642,3368938151,3353289242,47351,SRX21332627,SRS18578259,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94778,,0.14581,,0.80028,,0.52885,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25113,SRR25822230,SRX21332627,SRS18578259,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep3,GSM7702834,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702834,GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq,GSM7702834 r1,GSM7702834,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_pos_Mcherry_S3_L001_I1_002.fastq.gz 2707_pos_Mcherry_S3_L001_I2_002.fastq.gz 2707_pos_Mcherry_S3_L001_R1_002.fastq.gz 2707_pos_Mcherry_S3_L001_R2_002.fastq.gz,fastq fastq fastq fastq,20078121078.0,145493631.0,GSM7702834 r3,0:10 1:10 2:28 3:90,A:3634984131;C:2943891505;G:3255149272;T:3259243920;N:1157962,10,10,28,90,3634984131,2943891505,3255149272,3259243920,1157962,SRX21332627,SRS18578259,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94884,,0.14622,,0.80099,,0.52572,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25114,SRR25822231,SRX21332627,SRS18578259,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep3,GSM7702834,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep3,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702834,GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq,GSM7702834 r1,GSM7702834,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,2707_pos_Mcherry_S3_L002_I1_002.fastq.gz 2707_pos_Mcherry_S3_L002_I2_002.fastq.gz 2707_pos_Mcherry_S3_L002_R1_002.fastq.gz 2707_pos_Mcherry_S3_L002_R2_002.fastq.gz,fastq fastq fastq fastq,20884392978.0,151336181.0,GSM7702834 r4,0:10 1:10 2:28 3:90,A:3779370721;C:3059502508;G:3396295214;T:3383865877;N:1221970,10,10,28,90,3779370721,3059502508,3396295214,3383865877,1221970,SRX21332627,SRS18578259,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.94896,,0.14559,,0.8002,,0.51418,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25115,SRR25605436,SRX21332626,SRS18578258,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep2,GSM7702833,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702833,GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq,GSM7702833 r1,GSM7702833,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py|options:  allowEarlyFileEnd,1907_GFP_pos_S2_L001_I1_001.fastq.gz 1907_GFP_pos_S2_L001_I2_001.fastq.gz 1907_GFP_pos_S2_L001_R1_001.fastq.gz 1907_GFP_pos_S2_L001_R2_001.fastq.gz,fastq fastq fastq fastq,22174775364.0,160686778.0,GSM7702833 r1,0:10 1:10 2:28 3:90,A:4207510387;C:3050740898;G:3362227609;T:3839333241;N:1997885,10,10,28,90,4207510387,3050740898,3362227609,3839333241,1997885,SRX21332626,SRS18578258,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93287,,0.11844,,0.81087,,0.55264,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25116,SRR25605437,SRX21332626,SRS18578258,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep2,GSM7702833,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702833,GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq,GSM7702833 r1,GSM7702833,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py|options:  allowEarlyFileEnd,1907_GFP_pos_S2_L002_I1_001.fastq.gz 1907_GFP_pos_S2_L002_I2_001.fastq.gz 1907_GFP_pos_S2_L002_R1_001.fastq.gz 1907_GFP_pos_S2_L002_R2_001.fastq.gz,fastq fastq fastq fastq,22047847518.0,159767011.0,GSM7702833 r2,0:10 1:10 2:28 3:90,A:4174013532;C:3033959926;G:3357962867;T:3813042664;N:52001,10,10,28,90,4174013532,3033959926,3357962867,3813042664,52001,SRX21332626,SRS18578258,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93359,,0.11585,,0.81087,,0.55705,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25117,SRR25822228,SRX21332626,SRS18578258,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep2,GSM7702833,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702833,GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq,GSM7702833 r1,GSM7702833,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_GFP_pos_S2_L001_I1_002.fastq.gz 1907_GFP_pos_S2_L001_I2_002.fastq.gz 1907_GFP_pos_S2_L001_R1_002.fastq.gz 1907_GFP_pos_S2_L001_R2_002.fastq.gz,fastq fastq fastq fastq,22010894568.0,159499236.0,GSM7702833 r3,0:10 1:10 2:28 3:90,A:4176090595;C:3026940106;G:3336506731;T:3814128890;N:1264918,10,10,28,90,4176090595,3026940106,3336506731,3814128890,1264918,SRX21332626,SRS18578258,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93457,,0.11733,,0.81032,,0.55203,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25118,SRR25822229,SRX21332626,SRS18578258,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep2,GSM7702833,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702833,GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq,GSM7702833 r1,GSM7702833,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_GFP_pos_S2_L002_I1_002.fastq.gz 1907_GFP_pos_S2_L002_I2_002.fastq.gz 1907_GFP_pos_S2_L002_R1_002.fastq.gz 1907_GFP_pos_S2_L002_R2_002.fastq.gz,fastq fastq fastq fastq,22227220746.0,161066817.0,GSM7702833 r4,0:10 1:10 2:28 3:90,A:4209198596;C:3055507768;G:3382717459;T:3847294849;N:1294858,10,10,28,90,4209198596,3055507768,3382717459,3847294849,1294858,SRX21332626,SRS18578258,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93428,,0.11629,,0.81014,,0.56014,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25119,SRR25605438,SRX21332625,SRS18578257,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep2,GSM7702832,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702832,GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq,GSM7702832 r1,GSM7702832,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_pos_pos_S1_L001_I1_001.fastq.gz 1907_pos_pos_S1_L001_I2_001.fastq.gz 1907_pos_pos_S1_L001_R1_001.fastq.gz 1907_pos_pos_S1_L001_R2_001.fastq.gz,fastq fastq fastq fastq,25141955454.0,182188083.0,GSM7702832 r1,0:10 1:10 2:28 3:90,A:4942067104;C:3446689328;G:3778138998;T:4227670796;N:2361244,10,10,28,90,4942067104,3446689328,3778138998,4227670796,2361244,SRX21332625,SRS18578257,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.90803,,0.11273,,0.81872,,0.54345,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25120,SRR25605439,SRX21332625,SRS18578257,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep2,GSM7702832,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702832,GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq,GSM7702832 r1,GSM7702832,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_pos_pos_S1_L002_R2_001.fastq.gz 1907_pos_pos_S1_L002_R1_001.fastq.gz 1907_pos_pos_S1_L002_I2_001.fastq.gz 1907_pos_pos_S1_L002_I1_001.fastq.gz,fastq fastq fastq fastq,25433730786.0,184302397.0,GSM7702832 r2,0:10 1:10 2:28 3:90,A:4980326106;C:3488986366;G:3842594871;T:4275245265;N:63122,10,10,28,90,4980326106,3488986366,3842594871,4275245265,63122,SRX21332625,SRS18578257,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.91035,,0.11175,,0.82016,,0.52814,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25121,SRR25822226,SRX21332625,SRS18578257,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep2,GSM7702832,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702832,GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq,GSM7702832 r1,GSM7702832,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_pos_pos_S1_L001_R2_002.fastq.gz 1907_pos_pos_S1_L001_R1_002.fastq.gz 1907_pos_pos_S1_L001_I2_002.fastq.gz 1907_pos_pos_S1_L001_I1_002.fastq.gz,fastq fastq fastq fastq,24834656088.0,179961276.0,GSM7702832 r3,0:10 1:10 2:28 3:90,A:4878672093;C:3403443000;G:3732878174;T:4180051888;N:1469685,10,10,28,90,4878672093,3403443000,3732878174,4180051888,1469685,SRX21332625,SRS18578257,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.90776,,0.11325,,0.81852,,0.54011,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25122,SRR25822227,SRX21332625,SRS18578257,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep2,GSM7702832,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep2,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702832,GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq,GSM7702832 r1,GSM7702832,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,SINGLE,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,1907_pos_pos_S1_L002_I1_002.fastq.gz 1907_pos_pos_S1_L002_I2_002.fastq.gz 1907_pos_pos_S1_L002_R1_002.fastq.gz 1907_pos_pos_S1_L002_R2_002.fastq.gz,fastq fastq fastq fastq,25609437216.0,185575632.0,GSM7702832 r4,0:10 1:10 2:28 3:90,A:5015380345;C:3509305882;G:3867598814;T:4308011415;N:1510424,10,10,28,90,5015380345,3509305882,3867598814,4308011415,1510424,SRX21332625,SRS18578257,SRA1702612,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.90907,,0.11192,,0.81994,,0.50902,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25123,SRR25605440,SRX21332624,SRS18578256,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep1,GSM7702831,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep1,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702831,GSM7702831: gfp enriched rep1; Danio rerio; RNA Seq,GSM7702831 r1,GSM7702831,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,gfp_nre_S1_L001_R2_001.fastq.gz gfp_nre_S1_L001_R1_001.fastq.gz gfp_nre_S1_L001_I2_001.fastq.gz gfp_nre_S1_L001_I1_001.fastq.gz,fastq fastq fastq fastq,47873499684.0,346909418.0,GSM7702831 r1,0:10 1:10 2:28 3:90,A:8912593789;C:6786299583;G:7771099171;T:7737013838;N:14841239,10,10,28,90,8912593789,6786299583,7771099171,7737013838,14841239,SRX21332624,SRS18578256,SRA1690580,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.90717,,0.15854,,0.80501,,0.50927,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25124,SRR25605441,SRX21332624,SRS18578256,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,gfp enriched rep1,GSM7702831,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,gfp enriched rep1,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702831,GSM7702831: gfp enriched rep1; Danio rerio; RNA Seq,GSM7702831 r1,GSM7702831,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,gfp_nre_S1_L002_I1_001.fastq.gz gfp_nre_S1_L002_I2_001.fastq.gz gfp_nre_S1_L002_R1_001.fastq.gz gfp_nre_S1_L002_R2_001.fastq.gz,fastq fastq fastq fastq,48659347860.0,352603970.0,GSM7702831 r2,0:10 1:10 2:28 3:90,A:9056524463;C:6888387179;G:7922807308;T:7854042793;N:12595557,10,10,28,90,9056524463,6888387179,7922807308,7854042793,12595557,SRX21332624,SRS18578256,SRA1690580,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.90541,,0.15686,,0.80543,,0.51051,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25125,SRR25605442,SRX21332623,SRS18578255,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep1,GSM7702830,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep1,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702830,GSM7702830: mcherry enriched rep1; Danio rerio; RNA Seq,GSM7702830 r1,GSM7702830,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,double_pos_S2_L001_R2_001.fastq.gz double_pos_S2_L001_R1_001.fastq.gz double_pos_S2_L001_I2_001.fastq.gz double_pos_S2_L001_I1_001.fastq.gz,fastq fastq fastq fastq,41371188084.0,299791218.0,GSM7702830 r1,0:10 1:10 2:28 3:90,A:7690531012;C:5787287634;G:6514950407;T:6975618202;N:12822365,10,10,28,90,7690531012,5787287634,6514950407,6975618202,12822365,SRX21332623,SRS18578255,SRA1690580,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93894,,0.17677,,0.79926,,0.51122,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
25126,SRR25605443,SRX21332623,SRS18578255,SRP454539,PRJNA1004255,Unique activities of two overlapping PAX6 retinal enhancers,GSE240575,Transcriptome Analysis,Enhancers play a critical role in development by precisely modulating spatial  temporal  and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease  however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development  with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive  redundant  or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers  HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos  we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping  these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active  we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique  we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.,,pubmed:37643867,,mcherry enriched rep1,GSM7702830,,source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing,mcherry enriched rep1,Cell Ranger v6.1.2 was used to perform alignment  filtering  barcode counting  and UMI counting. A custom reference genome was created for alignment using cellranger mkref  combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al.  2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size  detected genes  and mitochondrial reads McCarthy et al.  2017. Cells with detected genes ≥ 500  library size ≥ 800  and mitochondrial reads ≤ 10% were retained. Within the processed dataset  mean reads per cell = 11239  and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al.  2019; Farnsworth et al.  2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types.  Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al.  2018. SCTransform was used to perform log normalisation  scaling  and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used  with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted to HVGs  using Seurat's RunPCA function with number of PCs = 50. To enable integration of the samples  we then used Harmony to generate PCs corrected for batch effects between libraries Korsunsky et al.  2019. The Harmony PCs were then used to perform K nearest neighbour analysis k=20 and Louvain clustering using Seurat 15 dimensions and resolution 0.6. Clusters were annotated as retinal cell types based on the highest expressed marker genes  and other known genes for each cell type  using information from the literature and ZFIN Sprague et al.  2008. Cell cycle scoring was performed using the Seurat CellCycleScoring function  using zebrafish genes homologous to the ‘s.features’ and ‘g2m.features’ genes provided by Seurat. Assembly: GRCz11 Supplementary files format and content: Tab separated values files  matrix files  Seurat object RDS file,Eye,,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.,,tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf,GSM7702830,GSM7702830: mcherry enriched rep1; Danio rerio; RNA Seq,GSM7702830 r1,GSM7702830,1,NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf  embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF  and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C  then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution  and once with FACSmax Amsbio. In a final 500 µl FACSmax  the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris  and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP454539,,loader:fastq load.py,double_pos_S2_L002_I1_001.fastq.gz double_pos_S2_L002_I2_001.fastq.gz double_pos_S2_L002_R1_001.fastq.gz double_pos_S2_L002_R2_001.fastq.gz,fastq fastq fastq fastq,42692812980.0,309368210.0,GSM7702830 r2,0:10 1:10 2:28 3:90,A:7933061913;C:5964617781;G:6744802923;T:7189579846;N:11076437,10,10,28,90,7933061913,5964617781,6744802923,7189579846,11076437,SRX21332623,SRS18578255,SRA1690580,"Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh","Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh",1,0.93887,,0.17578,,0.79825,,0.50169,,90,,B,,usable mapping rate,illumina,nextseq_v2,3prime,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United Kingdom,2023-08-10,Hatching,Embryo,Eye,Sensory System
26483,SRR25930975,SRX21649988,SRS18818460,SRP458853,PRJNA1013567,scRNA seq of 50 hpf and 80 hpf isolated zebrafish hearts,GSE242483,Transcriptome Analysis,Seeking to identify additional transcription factors required for cardiac valve formation  we determined and explored the transcriptional landscape of endocardial cells at the time when key morphogenetic events underlying valve development take place. Overall design: We isolated wild type zebrafish hearts at 50 hpf when valve identity has just been established  and at 80 hpf when forming valves are first observed,,pubmed:38748804,,50hpf,GSM7764481,,source name:heart|tissue:heart|genotype:wild type|age:50hpf|geo loc name:missing|collection date:missing,50hpf,Reads were aligned against the zebrafish genome and counted by StarSolo. Preprocessed counts were further analysed using Scanpy. Basic cell quality control was conducted by taking the number of detected genes and mitochondrial content into consideration. We removed cells that did not express more than 300 genes or had a mitochondrial content greater than 8%. Furthermore  we filtered genes if they were detected in less than 30 cells. Raw counts per cell were normalised to the median count over all cells and transformed into log space to stabilise variance. We initially reduced dimensionality of the dataset using PCA  retaining 50 principal components. Subsequent steps  like low dimensional UMAP embedding and cell clustering  were based on the initial PCA. Final data visualization was done by scanpy and cellxgene packages. Assembly: DanRer11 Supplementary files format and content: starsolo outputs,heart,,Whole hearts were isolated and cardiac cells were dissociated. Dead cells were removed from the final cell isolate by FACs sorting in PBS without xxx and Magnesium and with 0.04% BSA. Each sample was run separately on a lane in a Chromium controller with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3 1 10xGenomics.  scRNA seq library preparation was done using a standard protocol  and sequencing was done on a Nextseq2000,,tissue:heart|genotype:wild type|age:50hpf,GSM7764481,GSM7764481: 50hpf; Danio rerio; RNA Seq,GSM7764481 r1,GSM7764481,1,Whole hearts were isolated and cardiac cells were dissociated. Dead cells were removed from the final cell isolate by FACs sorting in PBS without xxx and Magnesium and with 0.04% BSA. Each sample was run separately on a lane in a Chromium controller with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3 1 10xGenomics.  scRNA seq library preparation was done using a standard protocol  and sequencing was done on a Nextseq2000,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP458853,,,Giulia_50hpf_S1_R2_001.fastq.gz Giulia_50hpf_S1_R1_001.fastq.gz,fastq fastq,19874763820.0,237988724.0,GSM7764481 r1,0:28 1:55.51,A:5320257914;C:4424267230;G:4320623789;T:5708249185;N:101365702,28,55,,,5320257914,4424267230,4320623789,5708249185,101365702,SRX21649988,SRS18818460,SRA1706674,"Bioinformatics, Max-Planck-Institute for Heart and Lung Research","Bioinformatics, Max-Planck-Institute for Heart and Lung Research",2,0.00185,0.94414,0.00087,0.13397,0.99675,0.80172,0.41142,0.51985,28,56,T,B,sc-like readlen,illumina,nextseq_v2,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,Germany,2023-09-06,Hatching,Embryo,Heart,Cardiovascular System
31872,SRR28764740,SRX24330059,SRS21091017,SRP502657,PRJNA1101966,Pioneer neurons are molecularly distinct  and their axon targeting is regulated by retinoic acid signaling,GSE264323,Transcriptome Analysis,During nervous system development  pioneer neurons are the first to extend their axons into target tissues  creating a scaffold for follower neurons. Despite years of study  whether pioneer neurons are a molecularly distinct population is unknown. Analysis of zebrafish posterior lateral line pLL sensory neurons during axon growth using single cell RNA sequencing scRNA seq revealed that pioneer and follower neurons are transcriptionally distinct. Expression profiling of differentiating pLL progenitors defined follower as the ground state  whereas “pioneer” is a later developmental state. The scRNA seq data revealed active retinoic acid RA signaling in followers  but not in pioneers. Modulation of RA signaling within single pLL neurons showed that its downregulation in pioneers is necessary for expression of neurotrophic factor receptor ret  which is required for correct targeting of pioneer axons. Our study provided insights into the molecular landscape of pioneer neurons and revealed the regulatory role of RA signaling in their development. Overall design: Fourteen hpf  eighteen hpf  twenty two hpf  forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 µl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 µm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 µl PBS/ 2% BSA in a siliconized 1.5mL tube.,,,,CEL210312AN WT 48h,GSM8222580,,source name:zebrafish embryo|cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf loc name:missing|collection date:missing,CEL210312AN WT 48h,using Cell Ranger version 3.1.0; 10X Genomics  Pleasanton  CA. USA Assembly: ZebraFishGRCz11 Supplementary files format and content: Tab separated values files and matrix files,zebrafish embryo,,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer’s solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer’s solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter’s instructions single cell 3’ v3 protocol  10x Genomics.,,cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf,GSM8222580,GSM8222580: CEL210312AN WT 48h; Danio rerio; RNA Seq,GSM8222580 r1,GSM8222580,1,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter's instructions single cell three prime v3 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP502657,,loader:fastq load.py,CEL210312AN_WT_48h_S100_L003_I1_001.fastq.gz CEL210312AN_WT_48h_S100_L003_R1_001.fastq.gz CEL210312AN_WT_48h_S100_L003_R2_001.fastq.gz,fastq fastq fastq,8597674702.0,67698226.0,GSM8222580 r1,0:8 1:28 2:91,A:1824246454;C:1278127991;G:1448333280;T:1609791598;N:39243,8,28,91,,1824246454,1278127991,1448333280,1609791598,39243,SRX24330059,SRS21091017,SRA1850361,Oregon Health and Science Univ,Oregon Health and Science Univ,,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2024-04-22,Hatching,Embryo,Embryo Imprecise,All anatomical structures
31873,SRR28764741,SRX24330059,SRS21091017,SRP502657,PRJNA1101966,Pioneer neurons are molecularly distinct  and their axon targeting is regulated by retinoic acid signaling,GSE264323,Transcriptome Analysis,During nervous system development  pioneer neurons are the first to extend their axons into target tissues  creating a scaffold for follower neurons. Despite years of study  whether pioneer neurons are a molecularly distinct population is unknown. Analysis of zebrafish posterior lateral line pLL sensory neurons during axon growth using single cell RNA sequencing scRNA seq revealed that pioneer and follower neurons are transcriptionally distinct. Expression profiling of differentiating pLL progenitors defined follower as the ground state  whereas “pioneer” is a later developmental state. The scRNA seq data revealed active retinoic acid RA signaling in followers  but not in pioneers. Modulation of RA signaling within single pLL neurons showed that its downregulation in pioneers is necessary for expression of neurotrophic factor receptor ret  which is required for correct targeting of pioneer axons. Our study provided insights into the molecular landscape of pioneer neurons and revealed the regulatory role of RA signaling in their development. Overall design: Fourteen hpf  eighteen hpf  twenty two hpf  forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 µl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 µm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 µl PBS/ 2% BSA in a siliconized 1.5mL tube.,,,,CEL210312AN WT 48h,GSM8222580,,source name:zebrafish embryo|cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf loc name:missing|collection date:missing,CEL210312AN WT 48h,using Cell Ranger version 3.1.0; 10X Genomics  Pleasanton  CA. USA Assembly: ZebraFishGRCz11 Supplementary files format and content: Tab separated values files and matrix files,zebrafish embryo,,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer’s solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer’s solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter’s instructions single cell 3’ v3 protocol  10x Genomics.,,cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf,GSM8222580,GSM8222580: CEL210312AN WT 48h; Danio rerio; RNA Seq,GSM8222580 r1,GSM8222580,1,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter's instructions single cell three prime v3 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP502657,,loader:fastq load.py,CEL210312AN_WT_48h_S101_L003_I1_001.fastq.gz CEL210312AN_WT_48h_S101_L003_R1_001.fastq.gz CEL210312AN_WT_48h_S101_L003_R2_001.fastq.gz,fastq fastq fastq,10521686856.0,82847928.0,GSM8222580 r2,0:8 1:28 2:91,A:2230570510;C:1564859730;G:1774018840;T:1969664072;N:48296,8,28,91,,2230570510,1564859730,1774018840,1969664072,48296,SRX24330059,SRS21091017,SRA1850361,Oregon Health and Science Univ,Oregon Health and Science Univ,,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2024-04-22,Hatching,Embryo,Embryo Imprecise,All anatomical structures
31874,SRR28764742,SRX24330059,SRS21091017,SRP502657,PRJNA1101966,Pioneer neurons are molecularly distinct  and their axon targeting is regulated by retinoic acid signaling,GSE264323,Transcriptome Analysis,During nervous system development  pioneer neurons are the first to extend their axons into target tissues  creating a scaffold for follower neurons. Despite years of study  whether pioneer neurons are a molecularly distinct population is unknown. Analysis of zebrafish posterior lateral line pLL sensory neurons during axon growth using single cell RNA sequencing scRNA seq revealed that pioneer and follower neurons are transcriptionally distinct. Expression profiling of differentiating pLL progenitors defined follower as the ground state  whereas “pioneer” is a later developmental state. The scRNA seq data revealed active retinoic acid RA signaling in followers  but not in pioneers. Modulation of RA signaling within single pLL neurons showed that its downregulation in pioneers is necessary for expression of neurotrophic factor receptor ret  which is required for correct targeting of pioneer axons. Our study provided insights into the molecular landscape of pioneer neurons and revealed the regulatory role of RA signaling in their development. Overall design: Fourteen hpf  eighteen hpf  twenty two hpf  forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 µl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 µm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 µl PBS/ 2% BSA in a siliconized 1.5mL tube.,,,,CEL210312AN WT 48h,GSM8222580,,source name:zebrafish embryo|cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf loc name:missing|collection date:missing,CEL210312AN WT 48h,using Cell Ranger version 3.1.0; 10X Genomics  Pleasanton  CA. USA Assembly: ZebraFishGRCz11 Supplementary files format and content: Tab separated values files and matrix files,zebrafish embryo,,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer’s solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer’s solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter’s instructions single cell 3’ v3 protocol  10x Genomics.,,cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf,GSM8222580,GSM8222580: CEL210312AN WT 48h; Danio rerio; RNA Seq,GSM8222580 r1,GSM8222580,1,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter's instructions single cell three prime v3 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP502657,,loader:fastq load.py,CEL210312AN_WT_48h_S102_L003_I1_001.fastq.gz CEL210312AN_WT_48h_S102_L003_R1_001.fastq.gz CEL210312AN_WT_48h_S102_L003_R2_001.fastq.gz,fastq fastq fastq,10393048429.0,81835027.0,GSM8222580 r3,0:8 1:28 2:91,A:2205422432;C:1544223539;G:1750760277;T:1946533138;N:48071,8,28,91,,2205422432,1544223539,1750760277,1946533138,48071,SRX24330059,SRS21091017,SRA1850361,Oregon Health and Science Univ,Oregon Health and Science Univ,,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2024-04-22,Hatching,Embryo,Embryo Imprecise,All anatomical structures
31875,SRR28764743,SRX24330059,SRS21091017,SRP502657,PRJNA1101966,Pioneer neurons are molecularly distinct  and their axon targeting is regulated by retinoic acid signaling,GSE264323,Transcriptome Analysis,During nervous system development  pioneer neurons are the first to extend their axons into target tissues  creating a scaffold for follower neurons. Despite years of study  whether pioneer neurons are a molecularly distinct population is unknown. Analysis of zebrafish posterior lateral line pLL sensory neurons during axon growth using single cell RNA sequencing scRNA seq revealed that pioneer and follower neurons are transcriptionally distinct. Expression profiling of differentiating pLL progenitors defined follower as the ground state  whereas “pioneer” is a later developmental state. The scRNA seq data revealed active retinoic acid RA signaling in followers  but not in pioneers. Modulation of RA signaling within single pLL neurons showed that its downregulation in pioneers is necessary for expression of neurotrophic factor receptor ret  which is required for correct targeting of pioneer axons. Our study provided insights into the molecular landscape of pioneer neurons and revealed the regulatory role of RA signaling in their development. Overall design: Fourteen hpf  eighteen hpf  twenty two hpf  forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 µl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 µm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 µl PBS/ 2% BSA in a siliconized 1.5mL tube.,,,,CEL210312AN WT 48h,GSM8222580,,source name:zebrafish embryo|cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf loc name:missing|collection date:missing,CEL210312AN WT 48h,using Cell Ranger version 3.1.0; 10X Genomics  Pleasanton  CA. USA Assembly: ZebraFishGRCz11 Supplementary files format and content: Tab separated values files and matrix files,zebrafish embryo,,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer’s solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer’s solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter’s instructions single cell 3’ v3 protocol  10x Genomics.,,cell type:neuron|tissue:zebrafish embryo|strain:mixed|age:48 hpf,GSM8222580,GSM8222580: CEL210312AN WT 48h; Danio rerio; RNA Seq,GSM8222580 r1,GSM8222580,1,Forty eight hpf TgBACneurod1:EGFPnl1 zebrafish embryos were collected and euthanized in 1.7 ml microcentrifuge tubes. Embryos were deyolked using a calcium free Ringer's solution 116 mM NaCl  2.6 mM KCl  5 mM HEPES pH 7.0  by gently pipetting up and down with a P200 pipet. Embryos were incubated for 5 minutes in Ringer's solution. Embryos were transferred to pre warmed protease solutions 0.25% trypsin  1 mM EDTA  pH 8.0  PBS and collagenase P/HBSS 100 mg/mL was added. Embryos were incubated at 28° C for 15 minutes and were homogenized every 5 minutes using a P1000 pipet. The Stop solution 6X  30% calf serum  6 mM CaCl2  PBS was added and samples were centrifuged 350xg  4° C for 5 minutes. Supernatant was removed and 1 mL of chilled suspension solution was added 1% FBS  0.8 mM CaCl2  50 U/mL penicillin  0.05 mg/mL streptomycin  DMEM. Samples were centrifuged again 350g  4° C for 5 minutes and supernatant was removed. 700 μl of chilled suspension solution was added and cells were resuspended by pipetting. Cells were passed through a 40 μm cell strainer into a FACs tube and kept on ice. GFP and RFP+ cells were FAC sorted on a BD Symphony cell sorter into sorting buffer 50 μl PBS/ 2% BSA in a siliconized 1.5mL tube. Library was performed according to the manufacter's instructions single cell three prime v3 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP502657,,loader:fastq load.py,CEL210312AN_WT_48h_S103_L003_I1_001.fastq.gz CEL210312AN_WT_48h_S103_L003_R1_001.fastq.gz CEL210312AN_WT_48h_S103_L003_R2_001.fastq.gz,fastq fastq fastq,1539235301.0,12119963.0,GSM8222580 r4,0:8 1:28 2:91,A:326058980;C:228846142;G:262343444;T:285660694;N:7373,8,28,91,,326058980,228846142,262343444,285660694,7373,SRX24330059,SRS21091017,SRA1850361,Oregon Health and Science Univ,Oregon Health and Science Univ,,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2024-04-22,Hatching,Embryo,Embryo Imprecise,All anatomical structures
32609,SRR29325925,SRX24842084,SRS21553259,SRP512516,PRJNA1121299,"Gene expression profile of single endocardial cells in 48 hpf and 72 hpf zebrafish embryos/larvae. In the study ""The heart is a niche for hematopoietic stem and progenitor cells in zebrafish.""",GSE269378,Transcriptome Analysis,We used single cell RNA expression to uncover the cellular and molecular heterogeneity of isolated endocardial cells in the heart. From this study  we uncovered the presence of an endocardial hematopoietic cluster  along with other significant clusters including valve endocardial and interstitial cells  and non valve cells. Overall design: Endocardial cells were isolated using FACS from physically extracted Tgkdrl:nls mCherry and Tgcd41:GFP hearts according to presence of mCherry positive cells  and further analyzed using scRNAseq.,,pubmed:39217144,,EC 2d,GSM8314110,,source name:Heart|tissue:Heart|cell type:Endocardial cells|genotype:Tgkdrl:nls mCherry; Tgcd41:GFP|developmental stage:48 hpf loc name:missing|collection date:missing,EC 2d,Sequencing was done on Nextseq2000 and raw reads were aligned against the zebrafish genome DanRer11 and counted by StarSolo Preprocessed counts were further analysed using Scanpy ollowed by secondary analysis in Annotated Data Format. Preprocessed counts were further analyzed using Scanpy. Basic cell quality control was conducted by taking the number of detected genes and mitochondrial content into consideration. We removed 5 cells in total that did not express more than 300 genes or had a mitochondrial content greater than 5%. Furthermore  we filtered 24259 genes if they were detected in less than 30 cells <0.01%. Raw counts per cell were normalized to the median count over all cells and transformed into log space to stabilize variance. We initially reduced dimensionality of the dataset using PCA  retaining 50 principal components. Subsequent steps  like low dimensional UMAP embedding McInnes & Healy  https://arxiv.org/abs/1802.03426 and cell clustering via community detection  were based on the initial PCA. Final data visualization was done by scanpy and cellxgene packages. Assembly: danRer11 Supplementary files format and content: starsolo outputs,Heart,,Embryos were raised in a solution composed of 0.33 Danieau's medium  containing the following concentrations: 17.4 mM NaCl  0.21 mM KCl  0.12 mM MgSO4·H2O  and 0.18 mM CaNO32  while being maintained at a stable temperature of 28°C. To isolate EdCs for the single cell RNA sequencing experiment  hearts from 48  and 72 hpf Tgkdrl:nls mCherry; Tgcd41:GFP larvae were manually dissected in DMEM + 10% FBS.   Hearts were then centrifuged for 60 seconds at 3000 rpm  washed with 1 mL Hanks’ Balanced Salt Solution and dissociated into single cells by incubating in 100 μl Enzyme 1 and 5 μl Enzyme 2 Pierce Cardiomyocyte Dissociation Kit  Thermo Fisher Scientific  Cat#88281 for 20 minutes at 300 rpm in a 30°C shaker 79.  The samples were centrifuged for 5 minutes at 3000 rpm  the supernatant was discarded  and fresh medium was added to the dissociated cells and passed through 40 μM filter polystyrene 5ml tubes.  Negative controls of non fluorescent hearts or single color fluorescent hearts were prepared to define the sorting gates.  Cells were sorted using the BD FACSAria™ III BD Biosciences instrument.   Live cells were selected by exclusion of DAPI using 30mW 405nm excitation paired with 450/50 nm band pass filter.  The software used for sorting and analysis is BD FACSDiva v8.0.1.  Single positive Tgkdrl:nls mCherry+ cells and double positive Tgkdrl:nls mCherry+; Tgcd41:GFP+ cells were sorted and collected into tubes with DMEM + 10% FBS.  Cells from both collected samples were immediately combined and processed for the single cell RNA sequencing.  mCherry fluorescence was measured with 50mW 561nm excitation paired with 610/20nm band pass filter. GFP fluorescence was measured with 50mW 488nm excitation paired with 530/30 band pass filter. The cell suspensions were counted with Moxi cell counter and diluted according to manufacturer’s protocol to obtain 4.000 2d HC and 8.000 3d  10.497 EC  1.569 HC single cell data points per sample  respectively. Each sample was run separately on a lane in Chromium controller with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 10xGenomics. Library preparation was conducted according to the manufacturer indications.,Embryos/larvae were grown in 28 degree incubator until extraction at 48 or 72 hpf,tissue:Heart|cell type:Endocardial cells|genotype:Tgkdrl:nls mCherry; Tgcd41:GFP|developmental stage:48 hpf,GSM8314110,GSM8314110: EC 2d; Danio rerio; RNA Seq,GSM8314110 r1,GSM8314110,1,Embryos were raised in a solution composed of 0.33 Danieau's medium  containing the following concentrations: 17.4 mM NaCl  0.21 mM KCl  0.12 mM MgSO4·H2O  and 0.18 mM CaNO32  while being maintained at a stable temperature of 28°C. To isolate EdCs for the single cell RNA sequencing experiment  hearts from 48  and 72 hpf Tgkdrl:nls mCherry; Tgcd41:GFP larvae were manually dissected in DMEM + 10% FBS.   Hearts were then centrifuged for 60 seconds at 3000 rpm  washed with 1 mL Hanks' Balanced Salt Solution and dissociated into single cells by incubating in 100 μl Enzyme 1 and 5 μl Enzyme 2 Pierce Cardiomyocyte Dissociation Kit  Thermo Fisher Scientific  Cat#88281 for 20 minutes at 300 rpm in a 30°C shaker 79.  The samples were centrifuged for 5 minutes at 3000 rpm  the supernatant was discarded  and fresh medium was added to the dissociated cells and passed through 40 μM filter polystyrene 5ml tubes.  Negative controls of non fluorescent hearts or single color fluorescent hearts were prepared to define the sorting gates.  Cells were sorted using the BD FACSAria™ III BD Biosciences instrument.   Live cells were selected by exclusion of DAPI using 30mW 405nm excitation paired with 450/50 nm band pass filter.  The software used for sorting and analysis is BD FACSDiva v8.0.1.  Single positive Tgkdrl:nls mCherry+ cells and double positive Tgkdrl:nls mCherry+; Tgcd41:GFP+ cells were sorted and collected into tubes with DMEM + 10% FBS.  Cells from both collected samples were immediately combined and processed for the single cell RNA sequencing.  mCherry fluorescence was measured with 50mW 561nm excitation paired with 610/20nm band pass filter. GFP fluorescence was measured with 50mW 488nm excitation paired with 530/30 band pass filter. The cell suspensions were counted with Moxi cell counter and diluted according to manufacturer's protocol to obtain 4.000 2d HC and 8.000 3d  10.497 EC  1.569 HC single cell data points per sample  respectively. Each sample was run separately on a lane in Chromium controller with Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 10xGenomics. Library preparation was conducted according to the manufacturer indications.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,NextSeq 2000,,SRP512516,,,Felix_10x_Lib_EC-2d_R1.fastq.gz Felix_10x_Lib_EC-2d_R2.fastq.gz,fastq fastq,37849382027.0,476135961.0,GSM8314110 r1,0:28 1:51.49,A:10212120425;C:8162504214;G:8573129268;T:10704929378;N:196698742,28,51,,,10212120425,8162504214,8573129268,10704929378,196698742,SRX24842084,SRS21553259,SRA1892449,MPI for heart and lung research,MPI for heart and lung research,,,,,,,,,,,,T,B,mate1 technical by mapping diff,illumina,nextseq_v2,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,Germany,2024-06-07,Hatching,Embryo,Heart,Cardiovascular System
32863,SRR30114185,SRX25586802,SRS22237546,SRP523914,PRJNA1126282,Danio rerio Single cell Transcriptome,PRJNA1126282,Transcriptome Analysis,Single cell sequencing of Zebrafish model with RFC1 mutation for loss of function study. Samples from zebrafishes at 2 or 4 dpf isolated from the whole head or whole brain respectively. Libraries prepared and sequenced with 10X Genomics three prime Chromium kits.,,,,WT sc 2dpf,WT sc 2dpf S1 L003,,strain:Tgrfc1+/+|age:2dpf|collection date:2024 02 19|geo loc name:Canada: Montreal|sex:N/A|tissue:Head|genotype:Wild Type|BioSampleModel:Model organism or animal,,,,,,,,,scRNA Seq of danio rerio: 2dpf head,WT sc 2dpf,WT sc 2dpf,Four brains or three heads per condition from 4dpf or 2dpf larvae respectively were transferred into 1.5ml tubes containing 250 l of papain solution 100l papain  100l DNase I and 200l L cysteine per 5 mL of DMEM/F12. The tissue was digested 20min at 37C with pipetting every 10min. Enzymatic digestion was stopped with washing solution 65l glucose 45 %  50L HEPES 1M and 0.5ml FBS qsp 10ml DPBS 1X and centrifuged at 800g for 5min at 4C. Supernatant was removed and pellet was resuspended in 50l of washing solution. The cell suspension was filtered with a cell strainer to remove aggregates. Libraries were prepared with a three prime Chromium kit and sent for 10X Genomics sequencing at a depth of 250 million reads 100bp ea from Illumina NovaSeq6000.,,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP523914,,,WT_sc_2dpf_S1_L003_R2.fastq.gz WT_sc_2dpf_S1_L003_R1.fastq.gz,fastq fastq,56807165786.0,281223593.0,WT sc 2dpf S1 L003 R1.fastq.gz,0:101 1:101,A:15279867415;C:9333139424;G:10011041111;T:22182483036;N:634800,101,101,,,15279867415,9333139424,10011041111,22182483036,634800,SRX25586802,SRS22237546,SRA1936077,CHUM Research Center|Neurosciences,CHUM Research Center,2,0.07599,0.93395,0.0191,0.17781,0.97636,0.81215,0.61682,0.56234,101,101,T,B,mate1 technical by mapping diff,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,Canada,2024-08-02,Hatching,Embryo,Head,Nervous System
32864,SRR30114186,SRX25586801,SRS22237545,SRP523914,PRJNA1126282,Danio rerio Single cell Transcriptome,PRJNA1126282,Transcriptome Analysis,Single cell sequencing of Zebrafish model with RFC1 mutation for loss of function study. Samples from zebrafishes at 2 or 4 dpf isolated from the whole head or whole brain respectively. Libraries prepared and sequenced with 10X Genomics three prime Chromium kits.,,,,RFC1 KO sc 2dpf,RFC1 KO sc 2dpf S2 L003,,strain:Tgrfc1 / |age:2dpf|collection date:2024 02 19|geo loc name:Canada: Montreal|sex:N/A|tissue:Head|genotype:RFC1 KO|BioSampleModel:Model organism or animal,,,,,,,,,scRNA Seq of danio rerio: 2dpf head,RFC1 KO sc 2dpf,RFC1 KO sc 2dpf,Four brains or three heads per condition from 4dpf or 2dpf larvae respectively were transferred into 1.5ml tubes containing 250 l of papain solution 100l papain  100l DNase I and 200l L cysteine per 5 mL of DMEM/F12. The tissue was digested 20min at 37C with pipetting every 10min. Enzymatic digestion was stopped with washing solution 65l glucose 45 %  50L HEPES 1M and 0.5ml FBS qsp 10ml DPBS 1X and centrifuged at 800g for 5min at 4C. Supernatant was removed and pellet was resuspended in 50l of washing solution. The cell suspension was filtered with a cell strainer to remove aggregates. Libraries were prepared with a three prime Chromium kit and sent for 10X Genomics sequencing at a depth of 250 million reads 100bp ea from Illumina NovaSeq6000.,,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP523914,,,RFC1-KO_sc_2dpf_S2_L003_R1.fastq.gz RFC1-KO_sc_2dpf_S2_L003_R2.fastq.gz,fastq fastq,45448109280.0,224990640.0,RFC1 KO sc 2dpf S2 L003 R1.fastq.gz,0:101 1:101,A:12090767157;C:7635667126;G:8209912637;T:17511260653;N:501707,101,101,,,12090767157,7635667126,8209912637,17511260653,501707,SRX25586801,SRS22237545,SRA1936077,CHUM Research Center|Neurosciences,CHUM Research Center,2,0.09244,0.93268,0.02271,0.19887,0.97544,0.80955,0.6375,0.56143,101,101,T,B,mate1 technical by mapping diff,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,Canada,2024-08-02,Hatching,Embryo,Head,Nervous System
33425,SRR30186680,SRX25652375,SRS22295177,SRP525362,PRJNA1146366,Spatiotemporal dynamics of the developing zebrafish enteric nervous system at the whole organ level,GSE274407,Transcriptome Analysis,Neural crest cells give rise to the neurons of the enteric nervous system ENS that innervate the gastrointestinal tract to regulate gut motility. The immense size and distinct subregions of the gut present a challenge to understanding the spatial organization and sequential differentiation of different neuronal subtypes. Here  we profile enteric neurons and progenitors at single cell resolution during zebrafish embryonic and larval development to provide a near complete picture of transcriptional changes that accompany emergence of ENS neurons throughout the gastrointestinal tract. Multiplex spatial RNA transcript analysis reveals the temporal order and distinct localization patterns of neuronal subtypes along the length of the gut. Finally  we show that functional perturbation of select transcription factors Ebf1a  Gata3 and Satb2 alters the cell fate choice  respectively  of inhibitory  excitatory and serotonergic neuronal subtypes in the developing ENS. Overall design: Cells in the developing ENS of the TgPhox2bb:mNeonGreen zebrafish were isolated by Fluorescence activated cell sorting FACS according to the presence of mNeonGreen signal and used for scRNA seq experiment.,,pubmed:39642879,,ENS2dpf,GSM8448582,,source name:intestine|tissue:intestine|cell type:Cells in the ENS|genotype:Phox2bb:mNeonGreen heterozygous|age:2dpf|geo loc name:missing|collection date:missing,ENS2dpf,The raw data from 5 different time points were preprocessed separately using Kallisto and bustools programs. We constructed a reference transcriptome for pseudoalignment based on zebrafish genome assembly GRCz10. Assembly: GRCz10 Supplementary files format and content: h5ad files,intestine,,To isolate ENS cells  we generated a reporter line by inserting mNeonGreen fluorescent protein post the Phox2bb coding sequence  such that mNeonGreen faithfully recapitulates endogenous Phox2bb expression. Embryos and larvae at 2  3  4  5 dpf and 6 dpf were anesthetized by tricaine Methanesulfonate MS222  followed by deyolking and dissection to remove the head part. The remainder of the embryo/larvae were kept in Ringer’s Solution on ice and dissociated with Accumax cell dissociation solution at 30oC for 20  23  28  32 and 36 mins at 1000rpm in the Eppendorf thermomixer R system  with pipetting up and down every 5 minutes. post digestion  Accumax was quenched by ice cold resuspension buffer 1X Hanks' Balanced Salt Solution  no calcium  no magnesium HBSS;10mM HEPES  adjusted to PH 8.0; 2.5 mg/ml BSA  and samples were then passed through a 70um cell strainer into 50ml conical tube  centrifuged at 300g for 10 mins and resuspended in 500ul fresh resuspension buffer for cell sorting. Library was performed following the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,tissue:intestine|cell type:Cells in the ENS|genotype:Phox2bb:mNeonGreen heterozygous|age:2dpf,GSM8448582,GSM8448582: ENS2dpf; Danio rerio; RNA Seq,GSM8448582 r1,GSM8448582,1,To isolate ENS cells  we generated a reporter line by inserting mNeonGreen fluorescent protein post the Phox2bb coding sequence  such that mNeonGreen faithfully recapitulates endogenous Phox2bb expression. Embryos and larvae at 2  3  4  5 dpf and 6 dpf were anesthetized by tricaine Methanesulfonate MS222  followed by deyolking and dissection to remove the head part. The remainder of the embryo/larvae were kept in Ringer's Solution on ice and dissociated with Accumax cell dissociation solution at 30oC for 20  23  28  32 and 36 mins at 1000rpm in the Eppendorf thermomixer R system  with pipetting up and down every 5 minutes. post digestion  Accumax was quenched by ice cold resuspension buffer 1X Hanks' Balanced Salt Solution  no calcium  no magnesium HBSS;10mM HEPES  adjusted to PH 8.0; 2.5 mg/ml BSA  and samples were then passed through a 70um cell strainer into 50ml conical tube  centrifuged at 300g for 10 mins and resuspended in 500ul fresh resuspension buffer for cell sorting. Library was performed following the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina HiSeq 4000,,SRP525362,,loader:fastq load.py,FT-SA26633_S1_L005_I1_001.fastq.gz FT-SA26633_S1_L005_R1_001.fastq.gz FT-SA26633_S1_L005_R2_001.fastq.gz,fastq fastq fastq,16641390240.0,53681904.0,GSM8448582 r1,0:8 1:151 2:151,A:3957785280;C:2573469243;G:2993166279;T:6685407270;N:2106936,8,151,151,,3957785280,2573469243,2993166279,6685407270,2106936,SRX25652375,SRS22295177,SRA1943746,"Bronner lab, Biology and Biological Engineering, Caltech","Bronner lab, Biology and Biological Engineering, Caltech",,,,,,,,,,,,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,random_priming,unknown,sc,single_cell_droplet,10x,,United States,2024-08-09,Hatching,Embryo,Gut,Digestive System
33426,SRR30186681,SRX25652375,SRS22295177,SRP525362,PRJNA1146366,Spatiotemporal dynamics of the developing zebrafish enteric nervous system at the whole organ level,GSE274407,Transcriptome Analysis,Neural crest cells give rise to the neurons of the enteric nervous system ENS that innervate the gastrointestinal tract to regulate gut motility. The immense size and distinct subregions of the gut present a challenge to understanding the spatial organization and sequential differentiation of different neuronal subtypes. Here  we profile enteric neurons and progenitors at single cell resolution during zebrafish embryonic and larval development to provide a near complete picture of transcriptional changes that accompany emergence of ENS neurons throughout the gastrointestinal tract. Multiplex spatial RNA transcript analysis reveals the temporal order and distinct localization patterns of neuronal subtypes along the length of the gut. Finally  we show that functional perturbation of select transcription factors Ebf1a  Gata3 and Satb2 alters the cell fate choice  respectively  of inhibitory  excitatory and serotonergic neuronal subtypes in the developing ENS. Overall design: Cells in the developing ENS of the TgPhox2bb:mNeonGreen zebrafish were isolated by Fluorescence activated cell sorting FACS according to the presence of mNeonGreen signal and used for scRNA seq experiment.,,pubmed:39642879,,ENS2dpf,GSM8448582,,source name:intestine|tissue:intestine|cell type:Cells in the ENS|genotype:Phox2bb:mNeonGreen heterozygous|age:2dpf|geo loc name:missing|collection date:missing,ENS2dpf,The raw data from 5 different time points were preprocessed separately using Kallisto and bustools programs. We constructed a reference transcriptome for pseudoalignment based on zebrafish genome assembly GRCz10. Assembly: GRCz10 Supplementary files format and content: h5ad files,intestine,,To isolate ENS cells  we generated a reporter line by inserting mNeonGreen fluorescent protein post the Phox2bb coding sequence  such that mNeonGreen faithfully recapitulates endogenous Phox2bb expression. Embryos and larvae at 2  3  4  5 dpf and 6 dpf were anesthetized by tricaine Methanesulfonate MS222  followed by deyolking and dissection to remove the head part. The remainder of the embryo/larvae were kept in Ringer’s Solution on ice and dissociated with Accumax cell dissociation solution at 30oC for 20  23  28  32 and 36 mins at 1000rpm in the Eppendorf thermomixer R system  with pipetting up and down every 5 minutes. post digestion  Accumax was quenched by ice cold resuspension buffer 1X Hanks' Balanced Salt Solution  no calcium  no magnesium HBSS;10mM HEPES  adjusted to PH 8.0; 2.5 mg/ml BSA  and samples were then passed through a 70um cell strainer into 50ml conical tube  centrifuged at 300g for 10 mins and resuspended in 500ul fresh resuspension buffer for cell sorting. Library was performed following the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,tissue:intestine|cell type:Cells in the ENS|genotype:Phox2bb:mNeonGreen heterozygous|age:2dpf,GSM8448582,GSM8448582: ENS2dpf; Danio rerio; RNA Seq,GSM8448582 r1,GSM8448582,1,To isolate ENS cells  we generated a reporter line by inserting mNeonGreen fluorescent protein post the Phox2bb coding sequence  such that mNeonGreen faithfully recapitulates endogenous Phox2bb expression. Embryos and larvae at 2  3  4  5 dpf and 6 dpf were anesthetized by tricaine Methanesulfonate MS222  followed by deyolking and dissection to remove the head part. The remainder of the embryo/larvae were kept in Ringer's Solution on ice and dissociated with Accumax cell dissociation solution at 30oC for 20  23  28  32 and 36 mins at 1000rpm in the Eppendorf thermomixer R system  with pipetting up and down every 5 minutes. post digestion  Accumax was quenched by ice cold resuspension buffer 1X Hanks' Balanced Salt Solution  no calcium  no magnesium HBSS;10mM HEPES  adjusted to PH 8.0; 2.5 mg/ml BSA  and samples were then passed through a 70um cell strainer into 50ml conical tube  centrifuged at 300g for 10 mins and resuspended in 500ul fresh resuspension buffer for cell sorting. Library was performed following the manufacter's instructions single cell three prime v2 protocol  10x Genomics.,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina HiSeq 4000,,SRP525362,,loader:fastq load.py,FT-SA26633_S1_L006_I1_001.fastq.gz FT-SA26633_S1_L006_R1_001.fastq.gz FT-SA26633_S1_L006_R2_001.fastq.gz,fastq fastq fastq,17304656010.0,55821471.0,GSM8448582 r2,0:8 1:151 2:151,A:4101777717;C:2763806899;G:3078134302;T:6911601509;N:2763815,8,151,151,,4101777717,2763806899,3078134302,6911601509,2763815,SRX25652375,SRS22295177,SRA1943746,"Bronner lab, Biology and Biological Engineering, Caltech","Bronner lab, Biology and Biological Engineering, Caltech",,,,,,,,,,,,T,B,mate1 technical by mapping diff,illumina,hiseq_era,unknown,random_priming,unknown,sc,single_cell_droplet,10x,,United States,2024-08-09,Hatching,Embryo,Gut,Digestive System
35385,SRR32649448,SRX27953870,SRS24319775,SRP569567,PRJNA1234533,Delta family protocadhins contribute to protoglomerular targeting of olfactory sensory neuron axons in the olfactory bulb,GSE291626,Transcriptome Analysis,To understand how neural circuits are assembled  it is essential to identify and characterize the axonal guidance cues and receptors that determine the axonal trajectories and connections between neurons. We performed single cell RNA sequencing of olfactory sensory neurons from zebrafish to identify candidate axonal guidance related genes that are differentially expressed according to sensory axon target location in the olfactory bulb. Among the candidates we identified were several members of the non clustered delta protocadherin family of adhesion molecules. We found that two members of the delta1 protocadherin family  pcdh7b and pcdh11  are most highly expressed in sensory neurons that project to the DZ protoglomerulus. Knocking down either one impairs the ability of sensory axons to terminate within the DZ protoglomerulus  but does not affect the ability of other sensory axons from terminating normally in the CZ protoglomerulus. In contrast  two members of the delta2 protocadherin family  pcdh10b and pcdh17  are most highly expressed in sensory neurons that project to the CZ protoglomerulus. Knocking down pcdh10b induces ectopic terminations of CZ projecting sensory axons. Knocking down pcdh17 induces substantial ectopic axonal trajectories and impairs CZ projecting sensory axons from finding and terminating in the CZ protoglomerulus. Knockdowns of either pcdh10b or pcdh17 do not affect DZ projecting sensory axons. These findings suggest that delta1 protocadherins help DZ projecting sensory axons enter and remain within the DZ protoglomerulus  while delta2 protocadherins help CZ projecting sensory axons navigate to the CZ protoglomerulus. Overall design: Roughly 2000 zebrafish embryos we sacraficed at 36  48 hpf and 72 hpf. Olfactory tissue was dissociated in FACSmax dissosciation medium and then FACsorted to isolate OMP:RFP positive neurons.,,,,OMP positive Olfactory Sensory Neurons from 48pf Zebrafish,GSM8838717,,source name:Olfactory Sensory Tissue|tissue:Olfactory Sensory Tissue|cell line:Olactory Sensory Neurons|cell type:Neurons|sample age:48 hpf|genotype:Tgomp:lyn RFP|geo loc name:missing|collection date:missing,OMP positive Olfactory Sensory Neurons from 48pf Zebrafish,Cell Ranger V3.0.1 Assembly: GRCz11 Supplementary files format and content: Demultiplexed 10x Genomics Files.,Olfactory Sensory Tissue,,10X Genomics V3.3,,tissue:Olfactory Sensory Tissue|cell line:Olactory Sensory Neurons|cell type:Neurons|sample age:48 hpf|genotype:Tgomp:lyn RFP,GSM8838717,GSM8838717: OMP positive Olfactory Sensory Neurons from 48pf Zebrafish; Danio rerio; RNA Seq,GSM8838717 r1,GSM8838717,1,10X Genomics V3.3,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP569567,,loader:fastq load.py,97364_S5_L001_I1_001.fastq.gz 97364_S5_L001_R1_001.fastq.gz 97364_S5_L001_R2_001.fastq.gz,fastq fastq fastq,7455743360.0,57351872.0,GSM8838717 r1,0:8 1:28 2:94,A:1588619193;C:1118423731;G:1232237797;T:1451670240;N:125007,8,28,94,,1588619193,1118423731,1232237797,1451670240,125007,SRX27953870,SRS24319775,SRA2091725,"Department of Neuroscience, University of Pennsylvania","Department of Neuroscience, University of Pennsylvania",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2025-03-11,Hatching,Embryo,Brain,Nervous System
35386,SRR32649449,SRX27953870,SRS24319775,SRP569567,PRJNA1234533,Delta family protocadhins contribute to protoglomerular targeting of olfactory sensory neuron axons in the olfactory bulb,GSE291626,Transcriptome Analysis,To understand how neural circuits are assembled  it is essential to identify and characterize the axonal guidance cues and receptors that determine the axonal trajectories and connections between neurons. We performed single cell RNA sequencing of olfactory sensory neurons from zebrafish to identify candidate axonal guidance related genes that are differentially expressed according to sensory axon target location in the olfactory bulb. Among the candidates we identified were several members of the non clustered delta protocadherin family of adhesion molecules. We found that two members of the delta1 protocadherin family  pcdh7b and pcdh11  are most highly expressed in sensory neurons that project to the DZ protoglomerulus. Knocking down either one impairs the ability of sensory axons to terminate within the DZ protoglomerulus  but does not affect the ability of other sensory axons from terminating normally in the CZ protoglomerulus. In contrast  two members of the delta2 protocadherin family  pcdh10b and pcdh17  are most highly expressed in sensory neurons that project to the CZ protoglomerulus. Knocking down pcdh10b induces ectopic terminations of CZ projecting sensory axons. Knocking down pcdh17 induces substantial ectopic axonal trajectories and impairs CZ projecting sensory axons from finding and terminating in the CZ protoglomerulus. Knockdowns of either pcdh10b or pcdh17 do not affect DZ projecting sensory axons. These findings suggest that delta1 protocadherins help DZ projecting sensory axons enter and remain within the DZ protoglomerulus  while delta2 protocadherins help CZ projecting sensory axons navigate to the CZ protoglomerulus. Overall design: Roughly 2000 zebrafish embryos we sacraficed at 36  48 hpf and 72 hpf. Olfactory tissue was dissociated in FACSmax dissosciation medium and then FACsorted to isolate OMP:RFP positive neurons.,,,,OMP positive Olfactory Sensory Neurons from 48pf Zebrafish,GSM8838717,,source name:Olfactory Sensory Tissue|tissue:Olfactory Sensory Tissue|cell line:Olactory Sensory Neurons|cell type:Neurons|sample age:48 hpf|genotype:Tgomp:lyn RFP|geo loc name:missing|collection date:missing,OMP positive Olfactory Sensory Neurons from 48pf Zebrafish,Cell Ranger V3.0.1 Assembly: GRCz11 Supplementary files format and content: Demultiplexed 10x Genomics Files.,Olfactory Sensory Tissue,,10X Genomics V3.3,,tissue:Olfactory Sensory Tissue|cell line:Olactory Sensory Neurons|cell type:Neurons|sample age:48 hpf|genotype:Tgomp:lyn RFP,GSM8838717,GSM8838717: OMP positive Olfactory Sensory Neurons from 48pf Zebrafish; Danio rerio; RNA Seq,GSM8838717 r1,GSM8838717,1,10X Genomics V3.3,,RNA-Seq,TRANSCRIPTOMIC SINGLE CELL,cDNA,PAIRED,ILLUMINA,Illumina NovaSeq 6000,,SRP569567,,loader:fastq load.py,97364_S5_L002_I1_001.fastq.gz 97364_S5_L002_R1_001.fastq.gz 97364_S5_L002_R2_001.fastq.gz,fastq fastq fastq,7445521460.0,57273242.0,GSM8838717 r2,0:8 1:28 2:94,A:1586585615;C:1115323543;G:1231863598;T:1449738354;N:173638,8,28,94,,1586585615,1115323543,1231863598,1449738354,173638,SRX27953870,SRS24319775,SRA2091725,"Department of Neuroscience, University of Pennsylvania","Department of Neuroscience, University of Pennsylvania",,,,,,,,,,,,B,,usable mapping rate,illumina,novaseq_era,unknown,cdna_unspecified,unknown,sc,single_cell_droplet,10x,,United States,2025-03-11,Hatching,Embryo,Brain,Nervous System
35567,SRR32928199,SRX28203119,SRS24552825,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.F01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:6|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 6,100,100,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.F01.fq.gz,fastq,800436.0,9529.0,expt2.broad.48.enzymatic.P01.F01.fq.gz,0:84,A:247920;C:168170;G:180799;T:203153;N:394,84,,,,247920,168170,180799,203153,394,SRX28203119,SRS24552825,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35568,SRR32928200,SRX28203118,SRS24552823,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.E01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:5|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 5,99,99,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.E01.fq.gz,fastq,496356.0,5909.0,expt2.broad.48.enzymatic.P01.E01.fq.gz,0:84,A:156600;C:99183;G:113282;T:127036;N:255,84,,,,156600,99183,113282,127036,255,SRX28203118,SRS24552823,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35569,SRR32928201,SRX28203117,SRS24552824,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.D01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:4|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 4,98,98,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.D01.fq.gz,fastq,2053800.0,24450.0,expt2.broad.48.enzymatic.P01.D01.fq.gz,0:84,A:668634;C:393806;G:431279;T:559022;N:1059,84,,,,668634,393806,431279,559022,1059,SRX28203117,SRS24552824,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35570,SRR32928202,SRX28203116,SRS24552821,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.C01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:3|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 3,97,97,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.C01.fq.gz,fastq,2159052.0,25703.0,expt2.broad.48.enzymatic.P01.C01.fq.gz,0:84,A:837784;C:348085;G:463530;T:508544;N:1109,84,,,,837784,348085,463530,508544,1109,SRX28203116,SRS24552821,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35571,SRR32928203,SRX28203115,SRS24552822,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.B01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:2|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 2,96,96,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.B01.fq.gz,fastq,3287508.0,39137.0,expt2.broad.48.enzymatic.P01.B01.fq.gz,0:84,A:1090202;C:644484;G:702124;T:848812;N:1886,84,,,,1090202,644484,702124,848812,1886,SRX28203115,SRS24552822,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35572,SRR32928204,SRX28203114,SRS24552819,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.enzymatic.P01.A01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:enzymatic|replicate:1|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  enzymatic dissociation  replicate 1,95,95,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.enzymatic.P01.A01.fq.gz,fastq,6623400.0,78850.0,expt2.broad.48.enzymatic.P01.A01.fq.gz,0:84,A:2915935;C:1012732;G:1256523;T:1434621;N:3589,84,,,,2915935,1012732,1256523,1434621,3589,SRX28203114,SRS24552819,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35573,SRR32928205,SRX28203113,SRS24552820,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.H02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:16|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 16,94,94,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.H02.fq.gz,fastq,18079152.0,215228.0,expt2.broad.48.bead.P18.H02.fq.gz,0:84,A:6013555;C:3409380;G:3853431;T:4793252;N:9534,84,,,,6013555,3409380,3853431,4793252,9534,SRX28203113,SRS24552820,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35574,SRR32928206,SRX28203112,SRS24552818,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.H01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:15|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 15,93,93,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.H01.fq.gz,fastq,13210764.0,157271.0,expt2.broad.48.bead.P18.H01.fq.gz,0:84,A:4207727;C:2577088;G:2821128;T:3597752;N:7069,84,,,,4207727,2577088,2821128,3597752,7069,SRX28203112,SRS24552818,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35575,SRR32928207,SRX28203111,SRS24552817,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.G02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:14|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 14,92,92,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.G02.fq.gz,fastq,14834736.0,176604.0,expt2.broad.48.bead.P18.G02.fq.gz,0:84,A:5041294;C:2753273;G:3108141;T:3924037;N:7991,84,,,,5041294,2753273,3108141,3924037,7991,SRX28203111,SRS24552817,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35603,SRR32928234,SRX28203083,SRS24552789,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.G01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:13|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 13,91,91,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.G01.fq.gz,fastq,14202468.0,169077.0,expt2.broad.48.bead.P18.G01.fq.gz,0:84,A:4692822;C:2685182;G:2975257;T:3841545;N:7662,84,,,,4692822,2685182,2975257,3841545,7662,SRX28203083,SRS24552789,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35605,SRR32928236,SRX28203081,SRS24552788,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.F02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:12|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 12,90,90,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.F02.fq.gz,fastq,16051728.0,191092.0,expt2.broad.48.bead.P18.F02.fq.gz,0:84,A:5221193;C:3079215;G:3420876;T:4321889;N:8555,84,,,,5221193,3079215,3420876,4321889,8555,SRX28203081,SRS24552788,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35606,SRR32928237,SRX28203080,SRS24552786,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.F01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:11|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 11,89,89,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.F01.fq.gz,fastq,19022388.0,226457.0,expt2.broad.48.bead.P18.F01.fq.gz,0:84,A:6375349;C:3646304;G:4032366;T:4958089;N:10280,84,,,,6375349,3646304,4032366,4958089,10280,SRX28203080,SRS24552786,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35607,SRR32928238,SRX28203079,SRS24552785,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.E02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:10|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 10,88,88,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.E02.fq.gz,fastq,17436720.0,207580.0,expt2.broad.48.bead.P18.E02.fq.gz,0:84,A:5912075;C:3260546;G:3682832;T:4572094;N:9173,84,,,,5912075,3260546,3682832,4572094,9173,SRX28203079,SRS24552785,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35608,SRR32928239,SRX28203078,SRS24552783,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.E01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:9|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 9,87,87,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.E01.fq.gz,fastq,12759768.0,151902.0,expt2.broad.48.bead.P18.E01.fq.gz,0:84,A:4143321;C:2424389;G:2695567;T:3489773;N:6718,84,,,,4143321,2424389,2695567,3489773,6718,SRX28203078,SRS24552783,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35609,SRR32928240,SRX28203077,SRS24552784,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.D02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:8|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 8,86,86,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.D02.fq.gz,fastq,17226132.0,205073.0,expt2.broad.48.bead.P18.D02.fq.gz,0:84,A:5722683;C:3229414;G:3698323;T:4566519;N:9193,84,,,,5722683,3229414,3698323,4566519,9193,SRX28203077,SRS24552784,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35610,SRR32928241,SRX28203076,SRS24552782,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.D01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:7|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 7,85,85,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.D01.fq.gz,fastq,14390124.0,171311.0,expt2.broad.48.bead.P18.D01.fq.gz,0:84,A:4584304;C:2823369;G:3092301;T:3882609;N:7541,84,,,,4584304,2823369,3092301,3882609,7541,SRX28203076,SRS24552782,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35611,SRR32928242,SRX28203075,SRS24552781,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.C02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:6|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 6,84,84,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.C02.fq.gz,fastq,18592560.0,221340.0,expt2.broad.48.bead.P18.C02.fq.gz,0:84,A:6370777;C:3468105;G:3988886;T:4754942;N:9850,84,,,,6370777,3468105,3988886,4754942,9850,SRX28203075,SRS24552781,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35612,SRR32928243,SRX28203074,SRS24552780,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.C01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:5|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 5,83,83,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.C01.fq.gz,fastq,12557076.0,149489.0,expt2.broad.48.bead.P18.C01.fq.gz,0:84,A:3977371;C:2443832;G:2711905;T:3417301;N:6667,84,,,,3977371,2443832,2711905,3417301,6667,SRX28203074,SRS24552780,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35613,SRR32928244,SRX28203073,SRS24552779,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.B02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:4|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 4,82,82,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.B02.fq.gz,fastq,14289828.0,170117.0,expt2.broad.48.bead.P18.B02.fq.gz,0:84,A:4795697;C:2698305;G:3083373;T:3704963;N:7490,84,,,,4795697,2698305,3083373,3704963,7490,SRX28203073,SRS24552779,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35614,SRR32928245,SRX28203072,SRS24552778,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.B01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:3|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 3,81,81,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.B01.fq.gz,fastq,6255144.0,74466.0,expt2.broad.48.bead.P18.B01.fq.gz,0:84,A:1953811;C:1213806;G:1335743;T:1748533;N:3251,84,,,,1953811,1213806,1335743,1748533,3251,SRX28203072,SRS24552778,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35616,SRR32928247,SRX28203070,SRS24552776,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.A02.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:2|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 2,80,80,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.A02.fq.gz,fastq,16441656.0,195734.0,expt2.broad.48.bead.P18.A02.fq.gz,0:84,A:5495147;C:3127531;G:3493145;T:4317176;N:8657,84,,,,5495147,3127531,3493145,4317176,8657,SRX28203070,SRS24552776,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35617,SRR32928248,SRX28203069,SRS24552775,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a large number of cells in order to determine the cell number and diversity captured by the method.,,expt2.broad.48.bead.P18.A01.fq.gz,,seq depth:broad many cells|Experiment id:2|dissociation method:bead|replicate:1|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  broad sequencing of many cells at low depth  48hpf  bead dissociation  replicate 1,79,79,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.broad.48.bead.P18.A01.fq.gz,fastq,14319564.0,170471.0,expt2.broad.48.bead.P18.A01.fq.gz,0:84,A:4614154;C:2749141;G:3043759;T:3904988;N:7522,84,,,,4614154,2749141,3043759,3904988,7522,SRX28203069,SRS24552775,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35644,SRR32928275,SRX28203042,SRS24552748,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.H01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:8|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 8,173,173,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.H01.fq.gz,fastq,10952424.0,130386.0,expt2.deep.48.enzymatic.P01.H01.fq.gz,0:84,A:3876382;C:2009479;G:2298766;T:2765953;N:1844,84,,,,3876382,2009479,2298766,2765953,1844,SRX28203042,SRS24552748,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35645,SRR32928276,SRX28203041,SRS24552747,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.G01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:7|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 7,172,172,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.G01.fq.gz,fastq,14085792.0,167688.0,expt2.deep.48.enzymatic.P01.G01.fq.gz,0:84,A:5224461;C:2624861;G:2935295;T:3298694;N:2481,84,,,,5224461,2624861,2935295,3298694,2481,SRX28203041,SRS24552747,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35646,SRR32928277,SRX28203040,SRS24552746,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.F01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:6|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 6,171,171,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.F01.fq.gz,fastq,9595404.0,114231.0,expt2.deep.48.enzymatic.P01.F01.fq.gz,0:84,A:3301852;C:1874682;G:2048715;T:2368498;N:1657,84,,,,3301852,1874682,2048715,2368498,1657,SRX28203040,SRS24552746,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35648,SRR32928279,SRX28203038,SRS24552744,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.E01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:5|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 5,170,170,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.E01.fq.gz,fastq,6317472.0,75208.0,expt2.deep.48.enzymatic.P01.E01.fq.gz,0:84,A:2175918;C:1173989;G:1412172;T:1554313;N:1080,84,,,,2175918,1173989,1412172,1554313,1080,SRX28203038,SRS24552744,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35649,SRR32928280,SRX28203037,SRS24552743,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.D01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:4|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 4,169,169,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.D01.fq.gz,fastq,26805408.0,319112.0,expt2.deep.48.enzymatic.P01.D01.fq.gz,0:84,A:9489671;C:4847582;G:5421907;T:7041421;N:4827,84,,,,9489671,4847582,5421907,7041421,4827,SRX28203037,SRS24552743,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35650,SRR32928281,SRX28203036,SRS24552742,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.C01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:3|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 3,168,168,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.C01.fq.gz,fastq,37386300.0,445075.0,expt2.deep.48.enzymatic.P01.C01.fq.gz,0:84,A:15837274;C:5306422;G:7863430;T:8372443;N:6731,84,,,,15837274,5306422,7863430,8372443,6731,SRX28203036,SRS24552742,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35728,SRR32928359,SRX28202958,SRS24552663,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.B01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:2|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 2,167,167,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.B01.fq.gz,fastq,43263192.0,515038.0,expt2.deep.48.enzymatic.P01.B01.fq.gz,0:84,A:15711103;C:7988920;G:8879458;T:10676082;N:7629,84,,,,15711103,7988920,8879458,10676082,7629,SRX28202958,SRS24552663,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35729,SRR32928361,SRX28202957,SRS24552664,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using enzymatic digestion. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.enzymatic.P01.A01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:enzymatic|replicate:1|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  enzymatic dissociation  replicate 1,166,166,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.enzymatic.P01.A01.fq.gz,fastq,111654984.0,1329226.0,expt2.deep.48.enzymatic.P01.A01.fq.gz,0:84,A:50939840;C:16257231;G:20762106;T:23675984;N:19823,84,,,,50939840,16257231,20762106,23675984,19823,SRX28202957,SRS24552664,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35730,SRR32928362,SRX28202956,SRS24552662,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.H02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:16|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 16,165,165,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.H02.fq.gz,fastq,238250880.0,2836320.0,expt2.deep.48.bead.P18.H02.fq.gz,0:84,A:87774764;C:41601134;G:48854415;T:59978555;N:42012,84,,,,87774764,41601134,48854415,59978555,42012,SRX28202956,SRS24552662,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35731,SRR32928363,SRX28202955,SRS24552661,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.H01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:15|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 15,164,164,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.H01.fq.gz,fastq,164680572.0,1960483.0,expt2.deep.48.bead.P18.H01.fq.gz,0:84,A:58086166;C:30009860;G:33521214;T:43034590;N:28742,84,,,,58086166,30009860,33521214,43034590,28742,SRX28202955,SRS24552661,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35732,SRR32928364,SRX28202954,SRS24552660,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.G02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:14|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 14,163,163,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.G02.fq.gz,fastq,198828420.0,2367005.0,expt2.deep.48.bead.P18.G02.fq.gz,0:84,A:74768071;C:34197749;G:40014604;T:49812638;N:35358,84,,,,74768071,34197749,40014604,49812638,35358,SRX28202954,SRS24552660,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35733,SRR32928365,SRX28202953,SRS24552658,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.G01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:13|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 13,162,162,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.G01.fq.gz,fastq,185114916.0,2203749.0,expt2.deep.48.bead.P18.G01.fq.gz,0:84,A:67685300;C:32656500;G:37099326;T:47641224;N:32566,84,,,,67685300,32656500,37099326,47641224,32566,SRX28202953,SRS24552658,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35734,SRR32928366,SRX28202952,SRS24552659,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.F02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:12|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 12,161,161,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.F02.fq.gz,fastq,204337224.0,2432586.0,expt2.deep.48.bead.P18.F02.fq.gz,0:84,A:73860885;C:36257985;G:41424562;T:52758013;N:35779,84,,,,73860885,36257985,41424562,52758013,35779,SRX28202952,SRS24552659,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35736,SRR32928368,SRX28202950,SRS24552656,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.F01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:11|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 11,160,160,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.F01.fq.gz,fastq,251949012.0,2999393.0,expt2.deep.48.bead.P18.F01.fq.gz,0:84,A:93851928;C:45256084;G:51250087;T:61546817;N:44096,84,,,,93851928,45256084,51250087,61546817,44096,SRX28202950,SRS24552656,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35737,SRR32928369,SRX28202949,SRS24552655,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.E02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:10|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 10,159,159,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.E02.fq.gz,fastq,232932168.0,2773002.0,expt2.deep.48.bead.P18.E02.fq.gz,0:84,A:87509680;C:40423993;G:47439421;T:57518203;N:40871,84,,,,87509680,40423993,47439421,57518203,40871,SRX28202949,SRS24552655,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35738,SRR32928370,SRX28202948,SRS24552654,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.E01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:9|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 9,158,158,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.E01.fq.gz,fastq,162318072.0,1932358.0,expt2.deep.48.bead.P18.E01.fq.gz,0:84,A:58032105;C:28749428;G:32874202;T:42634000;N:28337,84,,,,58032105,28749428,32874202,42634000,28337,SRX28202948,SRS24552654,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35739,SRR32928371,SRX28202947,SRS24552653,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.D02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:8|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 8,157,157,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.D02.fq.gz,fastq,227115504.0,2703756.0,expt2.deep.48.bead.P18.D02.fq.gz,0:84,A:83596590;C:39343820;G:47110602;T:57023984;N:40508,84,,,,83596590,39343820,47110602,57023984,40508,SRX28202947,SRS24552653,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35740,SRR32928372,SRX28202946,SRS24552652,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.D01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:7|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 7,156,156,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.D01.fq.gz,fastq,180858636.0,2153079.0,expt2.deep.48.bead.P18.D01.fq.gz,0:84,A:63790400;C:33184555;G:36908678;T:46943253;N:31750,84,,,,63790400,33184555,36908678,46943253,31750,SRX28202946,SRS24552652,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35741,SRR32928373,SRX28202945,SRS24552651,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.C02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:6|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 6,155,155,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.C02.fq.gz,fastq,254836176.0,3033764.0,expt2.deep.48.bead.P18.C02.fq.gz,0:84,A:96800715;C:44151366;G:53186596;T:60652370;N:45129,84,,,,96800715,44151366,53186596,60652370,45129,SRX28202945,SRS24552651,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35742,SRR32928374,SRX28202944,SRS24552649,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.C01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:5|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 5,154,154,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.C01.fq.gz,fastq,158669364.0,1888921.0,expt2.deep.48.bead.P18.C01.fq.gz,0:84,A:55554232;C:28711615;G:32786705;T:41589084;N:27728,84,,,,55554232,28711615,32786705,41589084,27728,SRX28202944,SRS24552649,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35743,SRR32928375,SRX28202943,SRS24552650,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.B02.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:4|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 4,153,153,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.B02.fq.gz,fastq,190474200.0,2267550.0,expt2.deep.48.bead.P18.B02.fq.gz,0:84,A:71029855;C:33202220;G:39545818;T:46662820;N:33487,84,,,,71029855,33202220,39545818,46662820,33487,SRX28202943,SRS24552650,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures
35744,SRR32928376,SRX28202942,SRS24552647,SRP575522,PRJNA1244443,An improved  high yield method for isolating nuclei from individual zebrafish embryos for single nucleus RNA sequencing,PRJNA1244443,Other,Zebrafish are an ideal system to study the effects of chemical  genetic  and environmental perturbations on development due to their high fecundity and fast growth. Recently  single cell sequencing has emerged as a powerful tool to measure the effect of these perturbations at a whole embryo scale. These types of experiments rely on the ability to isolate nuclei from a large number of individually barcoded zebrafish embryos in parallel. Here we report a method for efficiently isolating high quality nuclei from zebrafish embryos in a 96 well plate format by bead homogenization in a lysis buffer. Through head to head sciPlex RNA seq experiments  we demonstrate that this method represents a substantial improvement over enzymatic dissociation and that it is compatible with a wide range of developmental stages.,,,Sci plexRNAseq of a 48hpf whole zebrafish embryo dissociated using bead homogenization. This subpool was sequenced on a relatively smaller number of cells in order to determine the number of UMIs produced by the method.,,expt2.deep.48.bead.P18.B01.fq.gz,,seq depth:deep many umis|Experiment id:2|dissociation method:bead|replicate:3|strain:AB|isolate:not applicable|breed:not applicable|cultivar:not applicable|ecotype:not applicable|age:48 hpf|dev stage:48 hpf|collection date:2024 01 31|geo loc name:USA:Washington  Seattle|sex:not collected|tissue:Whole Embryo|sample type:Whole organism|BioSampleModel:Model organism or animal,,,,,,,,,sciPlex RNAseq of whole Danio rerio embryos: expt2  deep sequencing of limited cells  48hpf  bead dissociation  replicate 3,152,152,Libraries were generated using sciPlex RNAseq3 combinatorial indexing. Each fasta represents a sample split from a large multiplexed sequencing run. Cell IDs are in the header of each fastq.,,,OTHER,TRANSCRIPTOMIC SINGLE CELL,other,SINGLE,ILLUMINA,NextSeq 2000,,SRP575522,,,expt2.deep.48.bead.P18.B01.fq.gz,fastq,75929112.0,903918.0,expt2.deep.48.bead.P18.B01.fq.gz,0:84,A:26148298;C:13739979;G:15434234;T:20593156;N:13445,84,,,,26148298,13739979,15434234,20593156,13445,SRX28202942,SRS24552647,SRA2104467,University of Washington|Genome Sciences,University of Washington,,,,,,,,,,,,B,,usable mapping rate,illumina,nextseq_v2,unknown,other,unknown,sc,single_cell_generic,generic-scrnaseq-only,,United States,2025-03-31,Hatching,Embryo,Whole Organism,All anatomical structures