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85 rows where devstage_curation = "Hatching" and tissue_curation = "Eye"

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technology 3

experiment.library_source 2

experiment.library_selection 2

experiment.library_layout 2

tissue_curation_coarse 1

tissue_curation 1

  • Eye · 85 ✖

experiment.platform 1

experiment.library_strategy 1

devstage_curation_coarse 1

devstage_curation 1

  • Hatching · 85 ✖
Link rowid ▼ run.accession experiment.accession sample.accession study.accession bioproject study.title study.alias study.type study.abstract study.attributes study.PMIDs sample.description sample.title sample.alias sample.centername sample.attributes GEOsample.title GEOsample.dataprocessing GEOsample.source GEOsample.treatmentprotocol GEOsample.extractprotocol GEOsample.growthprotocol GEOsample.characteristics GEOsample.accession experiment.title experiment.alias experiment.library_name experiment.design_description experiment.library_construction_protocol experiment.attributes experiment.library_strategy experiment.library_source experiment.library_selection experiment.library_layout experiment.platform experiment.instrument_model experiment.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
19087 19087 ERR13834862 ERX13237628 ERS21098715 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48HC1 S20 R1 001.fastq.gz SAMEA116100635 GIMM ENA first public:2024 01 01|INSDC center name:GIMM|INSDC status:public|Submitter Id:48 HC1|collected by:Jaakko Lehtimaki|collection date:2021 11 24|common name:zebrafish|dev stage:48 hpf location country and/or sea:Portugal|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|sample name:48 HC1|scientific name:Danio rerio|tissue type:retina                 Raw reads: 48 HC1 webin reads 48 HC1         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 HC1 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48HC1_S20_R1_001.fastq.gz fastq 3768442091.0 38065677.0 webin reads 48 HC1 0:99.00 A:1055463738;C:792073627;G:817607736;T:1103223790;N:73200 99       1055463738 792073627 817607736 1103223790 73200 ERX13237628 ERS21098715 ERA30883416 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2024-01-01 Hatching Embryo Eye Sensory System
19088 19088 ERR13834951 ERX13237717 ERS21098721 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58PR3 S26 R1 001.fastq.gz 58 PR3   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 PR3 webin reads 58 PR3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 PR3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58PR3_S26_R1_001.fastq.gz fastq 3869469736.0 38850717.0 webin reads 58 PR3 0:99.60 A:1103909449;C:804146524;G:827344036;T:1134036010;N:33717 99       1103909449 804146524 827344036 1134036010 33717 ERX13237717 ERS21098721 ERA30883529 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19089 19089 ERR13835010 ERX13237776 ERS21098726 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58AC4 S31 R1 001.fastq.gz SAMEA116100646 GIMM ENA first public:2024 01 01|INSDC center name:GIMM|INSDC status:public|Submitter Id:58 AC4|collected by:Jaakko Lehtimaki|collection date:2021 12 02|common name:zebrafish|dev stage:58 hpf location country and/or sea:Portugal|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|sample name:58 AC4|scientific name:Danio rerio|tissue type:retina                 Raw reads: 58 AC4 webin reads 58 AC4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 AC4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58AC4_S31_R1_001.fastq.gz fastq 3607648651.0 36331933.0 webin reads 58 AC4 0:99.30 A:1038047870;C:736262498;G:759315202;T:1073962987;N:60094 99       1038047870 736262498 759315202 1073962987 60094 ERX13237776 ERS21098726 ERA30883721 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2024-01-01 Hatching Embryo Eye Sensory System
19090 19090 ERR13822794 ERX13225546 ERS21098708 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48PR2 S13 R1 001.fastq.gz 48 PR2   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 PR2 webin reads 48 PR2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 PR2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48PR2_S13_R1_001.fastq.gz fastq 3809299607.0 38404185.0 webin reads 48 PR2 0:99.19 A:1067728192;C:802585802;G:828435932;T:1110435904;N:113777 99       1067728192 802585802 828435932 1110435904 113777 ERX13225546 ERS21098708 ERA30879682 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19091 19091 ERR13834854 ERX13237620 ERS21098714 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48AC4 S19 R1 001.fastq.gz 48 AC4   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 AC4 webin reads 48 AC4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 AC4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48AC4_S19_R1_001.fastq.gz fastq 3410801894.0 34443068.0 webin reads 48 AC4 0:99.03 A:966160063;C:707754156;G:733039262;T:1003772756;N:75657 99       966160063 707754156 733039262 1003772756 75657 ERX13237620 ERS21098714 ERA30883390 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19092 19092 ERR13828824 ERX13231590 ERS21098710 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48PR4 S15 R1 001.fastq.gz 48 PR4   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 PR4 webin reads 48 PR4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 PR4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48PR4_S15_R1_001.fastq.gz fastq 4365943927.0 44278278.0 webin reads 48 PR4 0:98.60 A:1214448497;C:927512154;G:957491418;T:1266385355;N:106503 98       1214448497 927512154 957491418 1266385355 106503 ERX13231590 ERS21098710 ERA30883309 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19093 19093 ERR13834993 ERX13237759 ERS21098723 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58AC1 S28 R1 001.fastq.gz 58 AC1   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 AC1 webin reads 58 AC1         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 AC1 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58AC1_S28_R1_001.fastq.gz fastq 3710768269.0 37364763.0 webin reads 58 AC1 0:99.31 A:1060419097;C:761984557;G:786533722;T:1101603428;N:227465 99       1060419097 761984557 786533722 1101603428 227465 ERX13237759 ERS21098723 ERA30883659 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19094 19094 ERR13834875 ERX13237641 ERS21098717 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48HC3 S22 R1 001.fastq.gz 48 HC3   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 HC3 webin reads 48 HC3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 HC3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48HC3_S22_R1_001.fastq.gz fastq 3083078586.0 31073491.0 webin reads 48 HC3 0:99.22 A:856250957;C:655140994;G:677078911;T:894558582;N:49142 99       856250957 655140994 677078911 894558582 49142 ERX13237641 ERS21098717 ERA30883447 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19095 19095 ERR13834899 ERX13237665 ERS21098720 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58PR2 S25 R1 001.fastq.gz 58 PR2   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 PR2 webin reads 58 PR2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 PR2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58PR2_S25_R1_001.fastq.gz fastq 3849571818.0 38706841.0 webin reads 58 PR2 0:99.45 A:1089593011;C:803322469;G:827777017;T:1128836774;N:42547 99       1089593011 803322469 827777017 1128836774 42547 ERX13237665 ERS21098720 ERA30883518 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19096 19096 ERR13834889 ERX13237655 ERS21098719 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58PR1 S24 R1 001.fastq.gz 58 PR1   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 PR1 webin reads 58 PR1         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 PR1 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58PR1_S24_R1_001.fastq.gz fastq 3945671240.0 39514922.0 webin reads 58 PR1 0:99.85 A:1135075173;C:807489123;G:827964380;T:1175109762;N:32802 99       1135075173 807489123 827964380 1175109762 32802 ERX13237655 ERS21098719 ERA30883497 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19097 19097 ERR13835019 ERX13237785 ERS21098728 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58HC2 S33 R1 001.fastq.gz SAMEA116100648 GIMM ENA first public:2024 01 01|INSDC center name:GIMM|INSDC status:public|Submitter Id:58 HC2|collected by:Jaakko Lehtimaki|collection date:2021 12 02|common name:zebrafish|dev stage:58 hpf location country and/or sea:Portugal|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|sample name:58 HC2|scientific name:Danio rerio|tissue type:retina                 Raw reads: 58 HC2 webin reads 58 HC2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 HC2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58HC2_S33_R1_001.fastq.gz fastq 3711237069.0 37217240.0 webin reads 58 HC2 0:99.72 A:1070592288;C:757099665;G:780689044;T:1102575310;N:280762 99       1070592288 757099665 780689044 1102575310 280762 ERX13237785 ERS21098728 ERA30883748 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2024-01-01 Hatching Embryo Eye Sensory System
19100 19100 ERR13828836 ERX13231602 ERS21098712 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48AC2 S17 R1 001.fastq.gz 48 AC2   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 AC2 webin reads 48 AC2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 AC2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48AC2_S17_R1_001.fastq.gz fastq 3205206588.0 32424632.0 webin reads 48 AC2 0:98.85 A:902438127;C:672433401;G:694813369;T:935445436;N:76255 98       902438127 672433401 694813369 935445436 76255 ERX13231602 ERS21098712 ERA30883343 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19101 19101 ERR13835004 ERX13237770 ERS21098725 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58AC3 S30 R1 001.fastq.gz 58 AC3   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 AC3 webin reads 58 AC3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 AC3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58AC3_S30_R1_001.fastq.gz fastq 3782993133.0 38295959.0 webin reads 58 AC3 0:98.78 A:1080745150;C:779903026;G:803258485;T:1118994941;N:91531 98       1080745150 779903026 803258485 1118994941 91531 ERX13237770 ERS21098725 ERA30883697 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19102 19102 ERR13834868 ERX13237634 ERS21098716 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48HC2 S21 R1 001.fastq.gz SAMEA116100636 GIMM ENA first public:2024 01 01|INSDC center name:GIMM|INSDC status:public|Submitter Id:48 HC2|collected by:Jaakko Lehtimaki|collection date:2021 11 24|common name:zebrafish|dev stage:48 hpf location country and/or sea:Portugal|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|sample name:48 HC2|scientific name:Danio rerio|tissue type:retina                 Raw reads: 48 HC2 webin reads 48 HC2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 HC2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48HC2_S21_R1_001.fastq.gz fastq 3424347865.0 34510204.0 webin reads 48 HC2 0:99.23 A:959488173;C:721171249;G:745705870;T:997922014;N:60559 99       959488173 721171249 745705870 997922014 60559 ERX13237634 ERS21098716 ERA30883434 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2024-01-01 Hatching Embryo Eye Sensory System
19104 19104 ERR13834880 ERX13237646 ERS21098718 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48HC4 S23 R1 001.fastq.gz 48 HC4   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 HC4 webin reads 48 HC4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 HC4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48HC4_S23_R1_001.fastq.gz fastq 3660674315.0 36821474.0 webin reads 48 HC4 0:99.42 A:1035049987;C:763030378;G:788062167;T:1074303516;N:228267 99       1035049987 763030378 788062167 1074303516 228267 ERX13237646 ERS21098718 ERA30883470 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19105 19105 ERR13822867 ERX13225633 ERS21098709 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48PR3 S14 R1 001.fastq.gz 48 PR3   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 PR3 webin reads 48 PR3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 PR3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48PR3_S14_R1_001.fastq.gz fastq 4232170464.0 42534853.0 webin reads 48 PR3 0:99.50 A:1195251409;C:889346003;G:917240118;T:1230272855;N:60079 99       1195251409 889346003 917240118 1230272855 60079 ERX13225633 ERS21098709 ERA30879704 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19108 19108 ERR13828843 ERX13234350 ERS21098713 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48AC3 S18 R1 001.fastq.gz 48 AC3   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 AC3 webin reads 48 AC3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 AC3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48AC3_S18_R1_001.fastq.gz fastq 3733628911.0 37779462.0 webin reads 48 AC3 0:98.83 A:1047204450;C:786943605;G:812327274;T:1086893109;N:260473 98       1047204450 786943605 812327274 1086893109 260473 ERX13234350 ERS21098713 ERA30883366 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19113 19113 ERR13835032 ERX13237798 ERS21098730 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58HC4 S35 R1 001.fastq.gz 58 HC4   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 HC4 webin reads 58 HC4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 HC4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58HC4_S35_R1_001.fastq.gz fastq 4418556015.0 44331835.0 webin reads 58 HC4 0:99.67 A:1255080581;C:916490847;G:946212615;T:1300735054;N:36918 99       1255080581 916490847 946212615 1300735054 36918 ERX13237798 ERS21098730 ERA30883773 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19114 19114 ERR13828829 ERX13231595 ERS21098711 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 48AC1 S16 R1 001.fastq.gz 48 AC1   organism:Danio rerio|collection date:2021 11 24|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:48 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 48 AC1 webin reads 48 AC1         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 48 AC1 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 48AC1_S16_R1_001.fastq.gz fastq 3933669121.0 39695406.0 webin reads 48 AC1 0:99.10 A:1105258159;C:824538054;G:854253688;T:1149538152;N:81068 99       1105258159 824538054 854253688 1149538152 81068 ERX13231595 ERS21098711 ERA30883326 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19117 19117 ERR13835025 ERX13237791 ERS21098729 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58HC3 S34 R1 001.fastq.gz 58 HC3   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 HC3 webin reads 58 HC3         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 HC3 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58HC3_S34_R1_001.fastq.gz fastq 3523899212.0 35292613.0 webin reads 58 HC3 0:99.85 A:1012516999;C:723225071;G:745152410;T:1042976092;N:28640 99       1012516999 723225071 745152410 1042976092 28640 ERX13237791 ERS21098729 ERA30883757 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19118 19118 ERR13835014 ERX13237780 ERS21098727 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58HC1 S32 R1 001.fastq.gz 58 HC1   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 HC1 webin reads 58 HC1         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 HC1 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58HC1_S32_R1_001.fastq.gz fastq 3436493821.0 34597887.0 webin reads 58 HC1 0:99.33 A:980894291;C:707196785;G:729948858;T:1018400124;N:53763 99       980894291 707196785 729948858 1018400124 53763 ERX13237780 ERS21098727 ERA30883731 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19119 19119 ERR13834987 ERX13237753 ERS21098722 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58PR4 S27 R1 001.fastq.gz 58 PR4   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 PR4 webin reads 58 PR4         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 PR4 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58PR4_S27_R1_001.fastq.gz fastq 3069040814.0 30923422.0 webin reads 58 PR4 0:99.25 A:875776938;C:633951750;G:653308475;T:905949639;N:54012 99       875776938 633951750 653308475 905949639 54012 ERX13237753 ERS21098722 ERA30883641 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
19121 19121 ERR13834997 ERX13237763 ERS21098724 ERP164534 PRJEB80557 RNAseq of Danio Rerio Photoreceptors Amacrine and Horizontal cells collected 38 48 and 58 hpf e09e99a9-eabd-4a7d-8f92-2c4cf9f384ed Other This analysis was conducted to elucidate potential guidance of retinal horizontal cells HCs by photoreceptors PRs and/or amacrine cells ACs during the retinal lamination. We dissected retinae from three different developmental stages corresponding to different stages in HCs lamination process and separated HCs PRs and HCs based on their differential GFP and DsRed expression in the Lhx1:GFP; ptf1a:DsRed zebrafish transgenic line. Four replicates of each sample were lysed converted to cDNA sequenced using the SMART SEQ2 protocol. ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01   retinal neurons 58AC2 S29 R1 001.fastq.gz 58 AC2   organism:Danio rerio|collection date:2021 12 02|scientific name:Danio rerio|collected by:Jaakko Lehtimaki|common name:zebrafish|lab host:Tg 5.5ptf1a:DsRed; Tglhx1a:EGFP|dev stage:58 hpf type:retina|geographic location country and/or sea:Portugal                 Raw reads: 58 AC2 webin reads 58 AC2         RNA-Seq TRANSCRIPTOMIC RANDOM SINGLE ILLUMINA Illumina HiSeq X   ERP164534 Raw reads: 58 AC2 ENA FIRST PUBLIC:2025 01 01|ENA LAST UPDATE:2025 01 01 58AC2_S29_R1_001.fastq.gz fastq 3872162091.0 38898173.0 webin reads 58 AC2 0:99.55 A:1111663794;C:793747086;G:817730996;T:1148980297;N:39918 99       1111663794 793747086 817730996 1148980297 39918 ERX13237763 ERS21098724 ERA30883678 Gulbenkian Institute for Molecular Medicine|European Nucleotide Archive Gulbenkian Institute for Molecular Medicine                       B   usable mapping rate illumina hiseq_era unknown random_priming unknown sc single_cell_plate smartseq   Belgium 2025-01-01 Hatching Embryo Eye Sensory System
25107 25107 SRR25605432 SRX21332628 SRS18578260 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep3 GSM7702835   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702835 GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq GSM7702835 r1 GSM7702835 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_GFP_pos_S4_L001_R2_001.fastq.gz 2707_GFP_pos_S4_L001_R1_001.fastq.gz 2707_GFP_pos_S4_L001_I2_001.fastq.gz 2707_GFP_pos_S4_L001_I1_001.fastq.gz fastq fastq fastq fastq 21603309774.0 156545723.0 GSM7702835 r1 0:10 1:10 2:28 3:90 A:3960451458;C:3105338977;G:3480072824;T:3541218595;N:2033216 10 10 28 90 3960451458 3105338977 3480072824 3541218595 2033216 SRX21332628 SRS18578260 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94286   0.14175   0.78395   0.51541   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25108 25108 SRR25605433 SRX21332628 SRS18578260 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep3 GSM7702835   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702835 GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq GSM7702835 r1 GSM7702835 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_GFP_pos_S4_L002_I1_001.fastq.gz 2707_GFP_pos_S4_L002_I2_001.fastq.gz 2707_GFP_pos_S4_L002_R1_001.fastq.gz 2707_GFP_pos_S4_L002_R2_001.fastq.gz fastq fastq fastq fastq 22413465270.0 162416415.0 GSM7702835 r2 0:10 1:10 2:28 3:90 A:4104405257;C:3220235633;G:3625448968;T:3667329471;N:58021 10 10 28 90 4104405257 3220235633 3625448968 3667329471 58021 SRX21332628 SRS18578260 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94239   0.14234   0.78338   0.50616   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25109 25109 SRR25822232 SRX21332628 SRS18578260 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep3 GSM7702835   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702835 GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq GSM7702835 r1 GSM7702835 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_GFP_pos_S4_L001_R2_002.fastq.gz 2707_GFP_pos_S4_L001_R1_002.fastq.gz 2707_GFP_pos_S4_L001_I2_002.fastq.gz 2707_GFP_pos_S4_L001_I1_002.fastq.gz fastq fastq fastq fastq 21424338264.0 155248828.0 GSM7702835 r3 0:10 1:10 2:28 3:90 A:3928077020;C:3077672525;G:3451667186;T:3513734857;N:1242932 10 10 28 90 3928077020 3077672525 3451667186 3513734857 1242932 SRX21332628 SRS18578260 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.9433   0.14242   0.78328   0.52384   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25110 25110 SRR25822233 SRX21332628 SRS18578260 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep3 GSM7702835   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702835 GSM7702835: gfp enriched rep3; Danio rerio; RNA Seq GSM7702835 r1 GSM7702835 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_GFP_pos_S4_L002_I1_002.fastq.gz 2707_GFP_pos_S4_L002_I2_002.fastq.gz 2707_GFP_pos_S4_L002_R1_002.fastq.gz 2707_GFP_pos_S4_L002_R2_002.fastq.gz fastq fastq fastq fastq 22542479952.0 163351304.0 GSM7702835 r4 0:10 1:10 2:28 3:90 A:4130174287;C:3235362228;G:3643457599;T:3691295761;N:1327485 10 10 28 90 4130174287 3235362228 3643457599 3691295761 1327485 SRX21332628 SRS18578260 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94183   0.14124   0.78301   0.50517   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25111 25111 SRR25605434 SRX21332627 SRS18578259 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep3 GSM7702834   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702834 GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq GSM7702834 r1 GSM7702834 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_pos_Mcherry_S3_L001_R2_001.fastq.gz 2707_pos_Mcherry_S3_L001_R1_001.fastq.gz 2707_pos_Mcherry_S3_L001_I2_001.fastq.gz 2707_pos_Mcherry_S3_L001_I1_001.fastq.gz fastq fastq fastq fastq 20161522758.0 146097991.0 GSM7702834 r1 0:10 1:10 2:28 3:90 A:3649819934;C:2958286158;G:3267504300;T:3271384686;N:1824112 10 10 28 90 3649819934 2958286158 3267504300 3271384686 1824112 SRX21332627 SRS18578259 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94851   0.1483   0.80012   0.52549   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25112 25112 SRR25605435 SRX21332627 SRS18578259 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep3 GSM7702834   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702834 GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq GSM7702834 r1 GSM7702834 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_pos_Mcherry_S3_L002_I1_001.fastq.gz 2707_pos_Mcherry_S3_L002_I2_001.fastq.gz 2707_pos_Mcherry_S3_L002_R1_001.fastq.gz 2707_pos_Mcherry_S3_L002_R2_001.fastq.gz fastq fastq fastq fastq 20708273928.0 150059956.0 GSM7702834 r2 0:10 1:10 2:28 3:90 A:3746088654;C:3037032642;G:3368938151;T:3353289242;N:47351 10 10 28 90 3746088654 3037032642 3368938151 3353289242 47351 SRX21332627 SRS18578259 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94778   0.14581   0.80028   0.52885   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25113 25113 SRR25822230 SRX21332627 SRS18578259 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep3 GSM7702834   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702834 GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq GSM7702834 r1 GSM7702834 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_pos_Mcherry_S3_L001_I1_002.fastq.gz 2707_pos_Mcherry_S3_L001_I2_002.fastq.gz 2707_pos_Mcherry_S3_L001_R1_002.fastq.gz 2707_pos_Mcherry_S3_L001_R2_002.fastq.gz fastq fastq fastq fastq 20078121078.0 145493631.0 GSM7702834 r3 0:10 1:10 2:28 3:90 A:3634984131;C:2943891505;G:3255149272;T:3259243920;N:1157962 10 10 28 90 3634984131 2943891505 3255149272 3259243920 1157962 SRX21332627 SRS18578259 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94884   0.14622   0.80099   0.52572   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25114 25114 SRR25822231 SRX21332627 SRS18578259 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep3 GSM7702834   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep3 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702834 GSM7702834: mcherry enriched rep3; Danio rerio; RNA Seq GSM7702834 r1 GSM7702834 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 2707_pos_Mcherry_S3_L002_I1_002.fastq.gz 2707_pos_Mcherry_S3_L002_I2_002.fastq.gz 2707_pos_Mcherry_S3_L002_R1_002.fastq.gz 2707_pos_Mcherry_S3_L002_R2_002.fastq.gz fastq fastq fastq fastq 20884392978.0 151336181.0 GSM7702834 r4 0:10 1:10 2:28 3:90 A:3779370721;C:3059502508;G:3396295214;T:3383865877;N:1221970 10 10 28 90 3779370721 3059502508 3396295214 3383865877 1221970 SRX21332627 SRS18578259 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.94896   0.14559   0.8002   0.51418   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25115 25115 SRR25605436 SRX21332626 SRS18578258 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep2 GSM7702833   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702833 GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq GSM7702833 r1 GSM7702833 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py|options: allowEarlyFileEnd 1907_GFP_pos_S2_L001_I1_001.fastq.gz 1907_GFP_pos_S2_L001_I2_001.fastq.gz 1907_GFP_pos_S2_L001_R1_001.fastq.gz 1907_GFP_pos_S2_L001_R2_001.fastq.gz fastq fastq fastq fastq 22174775364.0 160686778.0 GSM7702833 r1 0:10 1:10 2:28 3:90 A:4207510387;C:3050740898;G:3362227609;T:3839333241;N:1997885 10 10 28 90 4207510387 3050740898 3362227609 3839333241 1997885 SRX21332626 SRS18578258 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.93287   0.11844   0.81087   0.55264   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25116 25116 SRR25605437 SRX21332626 SRS18578258 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep2 GSM7702833   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702833 GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq GSM7702833 r1 GSM7702833 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py|options: allowEarlyFileEnd 1907_GFP_pos_S2_L002_I1_001.fastq.gz 1907_GFP_pos_S2_L002_I2_001.fastq.gz 1907_GFP_pos_S2_L002_R1_001.fastq.gz 1907_GFP_pos_S2_L002_R2_001.fastq.gz fastq fastq fastq fastq 22047847518.0 159767011.0 GSM7702833 r2 0:10 1:10 2:28 3:90 A:4174013532;C:3033959926;G:3357962867;T:3813042664;N:52001 10 10 28 90 4174013532 3033959926 3357962867 3813042664 52001 SRX21332626 SRS18578258 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.93359   0.11585   0.81087   0.55705   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25117 25117 SRR25822228 SRX21332626 SRS18578258 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep2 GSM7702833   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702833 GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq GSM7702833 r1 GSM7702833 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_GFP_pos_S2_L001_I1_002.fastq.gz 1907_GFP_pos_S2_L001_I2_002.fastq.gz 1907_GFP_pos_S2_L001_R1_002.fastq.gz 1907_GFP_pos_S2_L001_R2_002.fastq.gz fastq fastq fastq fastq 22010894568.0 159499236.0 GSM7702833 r3 0:10 1:10 2:28 3:90 A:4176090595;C:3026940106;G:3336506731;T:3814128890;N:1264918 10 10 28 90 4176090595 3026940106 3336506731 3814128890 1264918 SRX21332626 SRS18578258 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.93457   0.11733   0.81032   0.55203   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25118 25118 SRR25822229 SRX21332626 SRS18578258 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   gfp enriched rep2 GSM7702833   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing gfp enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702833 GSM7702833: gfp enriched rep2; Danio rerio; RNA Seq GSM7702833 r1 GSM7702833 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_GFP_pos_S2_L002_I1_002.fastq.gz 1907_GFP_pos_S2_L002_I2_002.fastq.gz 1907_GFP_pos_S2_L002_R1_002.fastq.gz 1907_GFP_pos_S2_L002_R2_002.fastq.gz fastq fastq fastq fastq 22227220746.0 161066817.0 GSM7702833 r4 0:10 1:10 2:28 3:90 A:4209198596;C:3055507768;G:3382717459;T:3847294849;N:1294858 10 10 28 90 4209198596 3055507768 3382717459 3847294849 1294858 SRX21332626 SRS18578258 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.93428   0.11629   0.81014   0.56014   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25119 25119 SRR25605438 SRX21332625 SRS18578257 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep2 GSM7702832   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702832 GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq GSM7702832 r1 GSM7702832 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_pos_pos_S1_L001_I1_001.fastq.gz 1907_pos_pos_S1_L001_I2_001.fastq.gz 1907_pos_pos_S1_L001_R1_001.fastq.gz 1907_pos_pos_S1_L001_R2_001.fastq.gz fastq fastq fastq fastq 25141955454.0 182188083.0 GSM7702832 r1 0:10 1:10 2:28 3:90 A:4942067104;C:3446689328;G:3778138998;T:4227670796;N:2361244 10 10 28 90 4942067104 3446689328 3778138998 4227670796 2361244 SRX21332625 SRS18578257 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.90803   0.11273   0.81872   0.54345   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25120 25120 SRR25605439 SRX21332625 SRS18578257 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep2 GSM7702832   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702832 GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq GSM7702832 r1 GSM7702832 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_pos_pos_S1_L002_R2_001.fastq.gz 1907_pos_pos_S1_L002_R1_001.fastq.gz 1907_pos_pos_S1_L002_I2_001.fastq.gz 1907_pos_pos_S1_L002_I1_001.fastq.gz fastq fastq fastq fastq 25433730786.0 184302397.0 GSM7702832 r2 0:10 1:10 2:28 3:90 A:4980326106;C:3488986366;G:3842594871;T:4275245265;N:63122 10 10 28 90 4980326106 3488986366 3842594871 4275245265 63122 SRX21332625 SRS18578257 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.91035   0.11175   0.82016   0.52814   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25121 25121 SRR25822226 SRX21332625 SRS18578257 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep2 GSM7702832   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702832 GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq GSM7702832 r1 GSM7702832 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_pos_pos_S1_L001_R2_002.fastq.gz 1907_pos_pos_S1_L001_R1_002.fastq.gz 1907_pos_pos_S1_L001_I2_002.fastq.gz 1907_pos_pos_S1_L001_I1_002.fastq.gz fastq fastq fastq fastq 24834656088.0 179961276.0 GSM7702832 r3 0:10 1:10 2:28 3:90 A:4878672093;C:3403443000;G:3732878174;T:4180051888;N:1469685 10 10 28 90 4878672093 3403443000 3732878174 4180051888 1469685 SRX21332625 SRS18578257 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.90776   0.11325   0.81852   0.54011   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25122 25122 SRR25822227 SRX21332625 SRS18578257 SRP454539 PRJNA1004255 Unique activities of two overlapping PAX6 retinal enhancers GSE240575 Transcriptome Analysis Enhancers play a critical role in development by precisely modulating spatial temporal and cell type specific gene expression. Sequence variants in enhancers have been implicated in disease however establishing the functional consequences of these variants is challenging due to a lack of understanding of precise cell types and developmental stages where the enhancers are normally active. PAX6 is the master regulator of eye development with a regulatory landscape containing multiple enhancers driving expression in the eye. Whether these enhancers perform additive redundant or distinct functions is unknown. Here we describe the precise cell types and regulatory activity of two PAX6 retinal enhancers HS5 and NRE. Using a unique combination of live imaging and single cell RNA sequencing in dual enhancer reporter zebrafish embryos we uncover differences in the spatiotemporal activity of these enhancers. Our results show that although overlapping these enhancers have distinct activities in different cell types and therefore likely non redundant functions. This work demonstrates that unique cell type specific activities can be uncovered for apparently similar enhancers when investigated at high resolution in vivo. Overall design: In order to define the precise cell types within the retina where the PAX6 enhancers HS5 and NRE are active we carried out scRNA seq on eyes from NRE eGFP/HS5 mCherry zebrafish reporter embryos. With this technique we aimed to uncover cell type or transcriptional differences between the two enhancer active populations. We dissected eyes from 48 hpf NRE eGFP/HS5 mCherry embryos and used FACS to enrich for either mCherry positive/HS5 active cells or eGFP positive/NRE active cells. Three samples for each population were processed for scRNA seq using the 10x Genomics Chromium single cell three prime gene expression technology.   pubmed:37643867   mcherry enriched rep2 GSM7702832   source name:Eye|tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf|geo loc name:missing|collection date:missing mcherry enriched rep2 Cell Ranger v6.1.2 was used to perform alignment filtering barcode counting and UMI counting. A custom reference genome was created for alignment using cellranger mkref combining the Danio rerio GRCz11 genome assembly with manually annotated eGFP and mCherry sequences. Cell calling and QC: The emptyDrops function from DropletUtils was used to filter out empty droplets/barcodes not corresponding to cells Lun et al. 2019. Mitochondrial and ribosomal genes were excluded from the emptyDrops analysis to improve the filtering of droplets containing ambient RNA or cell fragments. The scater package was used to filter cells based on the QC metrics of library size detected genes and mitochondrial reads McCarthy et al. 2017. Cells with detected genes ≥ 500 library size ≥ 800 and mitochondrial reads ≤ 10% were retained. Within the processed dataset mean reads per cell = 11239 and median genes per cell = 1554. Reference mapping and filtering: The SingleR package was used to annotate cell types based on mapping to the zebrafish single cell transcriptome atlas Aran et al. 2019; Farnsworth et al. 2019. Expression matrix and cell annotation data were downloaded from the UCSC cell browser http://zebrafish dev.cells.ucsc.edu; only the 2 dpf data were used for mapping. Erroneously sorted cells of non retinal identity for example pigmented cell types such as melanocytes with high autofluorescence were filtered out at this stage. This was carried out to improve the resolution of clustering for retinal cell types. Clustering and cell type annotation: Seurat v4 was used for clustering and further analysis for a total of 6 288 cells Butler et al. 2018. SCTransform was used to perform log normalisation scaling and highly variable gene HVG detection on a dataset consisting of the 6 samples merged into one. Standard SCTransform options were used with regression of mitochondrial expression and cell cycle stage using ‘vars.to.regress’. We performed Principle Component Analysis PCA on the normalized counts matrix restricted… Eye   NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell 3’ gene expression technology v3.1.   tissue:Eye|genotype:NRE eGFP/HS5 mCherry|developmental stage:48 hpf GSM7702832 GSM7702832: mcherry enriched rep2; Danio rerio; RNA Seq GSM7702832 r1 GSM7702832 1 NRE eGFP/HS5 mCherry embryos were collected and treated with PTU from 12 hpf. At 48 hpf embryos were anaesthetised with Tricaine 20–30 mg/l and placed into Danieau's solution. Eyes were dissected from 100 150 embryos using fine forceps Dumont #5SF and immediately placed into Danieau's solution on ice. Samples were centrifuged at 300g for 1 minute at 4°C then washed with Danieau's solution. Washing step was carried out three times with Danieau's solution and once with FACSmax Amsbio. In a final 500 µl FACSmax the samples were passed through a 35 µm cell strainer to obtain single cell suspension on ice. Samples were sorted for mCherry and eGFP fluorescence using a FACS Aria II BD or CytoFLEX SRT Beckman Coulter machine. Forward and side scatter sorting was used to select single cells from clumps and debris and DAPI staining was used to exclude dead cells. Libraries were prepared using the 10x Genomics Chromium single cell three prime gene expression technology v3.1.   RNA-Seq TRANSCRIPTOMIC SINGLE CELL cDNA SINGLE ILLUMINA NextSeq 2000   SRP454539   loader:fastq load.py 1907_pos_pos_S1_L002_I1_002.fastq.gz 1907_pos_pos_S1_L002_I2_002.fastq.gz 1907_pos_pos_S1_L002_R1_002.fastq.gz 1907_pos_pos_S1_L002_R2_002.fastq.gz fastq fastq fastq fastq 25609437216.0 185575632.0 GSM7702832 r4 0:10 1:10 2:28 3:90 A:5015380345;C:3509305882;G:3867598814;T:4308011415;N:1510424 10 10 28 90 5015380345 3509305882 3867598814 4308011415 1510424 SRX21332625 SRS18578257 SRA1702612 Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh Wendy Bickmore, MRC Human Genetics Unit, University of Edinburgh 1 0.90907   0.11192   0.81994   0.50902   90   B   usable mapping rate illumina nextseq_v2 3prime cdna_unspecified unknown sc single_cell_droplet 10x   United Kingdom 2023-08-10 Hatching Embryo Eye Sensory System
25123 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… 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 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… 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 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… 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 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… 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
36381 36381 SRR514031 SRX155011 SRS346297 SRP013815 PRJNA168635 Id2a knockdown in zebrafish retina GSE38786 Transcriptome Analysis During vertebrate retinogenesis the precise balance between retinoblast proliferation and differentiation is spatially and temporally regulated through a number of intrinsic factors and extrinsic signaling pathways. Moreover there are complex gene regulatory network interactions between these intrinsic factors and extrinsic pathways which ultimately function to determine when retinoblasts exit the cell cycle and terminally differentiate. We recently uncovered a cell non autonomous role for the intrinsic HLH factor Id2a in regulating retinoblast proliferation and differentiation with Id2a deficient retinae containing an abundance of proliferative retinoblasts and an absence of terminally differentiated retinal neurons and glia. Here we report that Id2a function is necessary and sufficient to limit Notch pathway activity during retinogenesis. Id2a deficient retinae possess elevated levels of Notch pathway component gene expression while retinae overexpressing id2a possess reduced expression of Notch pathway component genes. Attenuation of Notch signaling activity by DAPT or by morpholino knockdown of Notch1a is sufficient to rescue both the proliferative and differentiation defects in Id2a deficient retinae. In addition to regulating Notch pathway activity through an RNA Seq and differential gene expression analysis of Id2a deficient retinae we identify a number of additional intrinsic and extrinsic regulatory pathway components whose expression is regulated by Id2a. These data highlight the integral role played by Id2a in the gene regulatory network governing the transition from retinoblast proliferation to terminal differentiation during vertebrate retinogenesis. Overall design: Two biological replicates for both Id2aMM and Id2aMO samples   pubmed:22981606   Zebrafish retina with id2a morpholino treatment replicate 2 GSM949497   source name:Zebrafish retina with id2a morpholino treatment replicate 2|tissue:retina|genotype/variation:id2a knockdown Zebrafish retina with id2a morpholino treatment replicate 2 Filter out all reads with no call ‘N’ and low complexity reads without xxx 4 bases. Take reads that both paired end reads are passed the above condition. Map to EnsEMBL cDNA sequence then calculate RPK100M read counts per kb per 100 million of reads Summarize mapped reads per each cDNA. Determine differentially expressed genes with DESeq Genome build: EnsEMBL version 66 cDNA sequences Supplementary files format and content: tab delimited text files include RPKM values for each Sample DESeq output for differentially expressed genes. Zebrafish retina with id2a morpholino treatment replicate 2   Total RNA was isolated from dissected retinae at 48 hpf using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 microgram of total retinal RNA per condition to generate cDNA libraries by poly A selection.   tissue:retina|genotype/variation:id2a knockdown GSM949497 GSM949497: Zebrafish retina with id2a morpholino treatment replicate 2; Danio rerio; RNA Seq GSM949497 2 GSM949497: Zebrafish retina with id2a morpholino treatment replicate 2 1   GEO Accession:GSM949497 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>202</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC><READ_SPEC><READ_INDEX>1</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Reverse</READ_TYPE><BASE_COORD>102</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> SRP013815     Uribe201204_id2aKD2.untie_2.fastq.gz fastq 12244707932.0 60617366.0 GSM949497 r1 0:101 1:101 A:3345422426;C:2833723441;G:2860980232;T:3204581833;N:0 101 101     3345422426 2833723441 2860980232 3204581833 0 SRX155011 SRS346297 SRA054377 GEO Department of Biomedical Engineering, Ulsan National Institute of Science and Technology 2 0.8674 0.87016 0.09985 0.10059 0.75442 0.75187 0.47467 0.47364 101 101 B B biological fallback assumption illumina hiseq_era unknown poly_a trueseq bulk unknown unknown   South Korea 2012-06-18 Hatching Embryo Eye Sensory System
36382 36382 SRR514030 SRX155010 SRS346296 SRP013815 PRJNA168635 Id2a knockdown in zebrafish retina GSE38786 Transcriptome Analysis During vertebrate retinogenesis the precise balance between retinoblast proliferation and differentiation is spatially and temporally regulated through a number of intrinsic factors and extrinsic signaling pathways. Moreover there are complex gene regulatory network interactions between these intrinsic factors and extrinsic pathways which ultimately function to determine when retinoblasts exit the cell cycle and terminally differentiate. We recently uncovered a cell non autonomous role for the intrinsic HLH factor Id2a in regulating retinoblast proliferation and differentiation with Id2a deficient retinae containing an abundance of proliferative retinoblasts and an absence of terminally differentiated retinal neurons and glia. Here we report that Id2a function is necessary and sufficient to limit Notch pathway activity during retinogenesis. Id2a deficient retinae possess elevated levels of Notch pathway component gene expression while retinae overexpressing id2a possess reduced expression of Notch pathway component genes. Attenuation of Notch signaling activity by DAPT or by morpholino knockdown of Notch1a is sufficient to rescue both the proliferative and differentiation defects in Id2a deficient retinae. In addition to regulating Notch pathway activity through an RNA Seq and differential gene expression analysis of Id2a deficient retinae we identify a number of additional intrinsic and extrinsic regulatory pathway components whose expression is regulated by Id2a. These data highlight the integral role played by Id2a in the gene regulatory network governing the transition from retinoblast proliferation to terminal differentiation during vertebrate retinogenesis. Overall design: Two biological replicates for both Id2aMM and Id2aMO samples   pubmed:22981606   Zebrafish retina with id2a morpholino treatment replicate 1 GSM949496   source name:Zebrafish retina with id2a morpholino treatment replicate 1|tissue:retina|genotype/variation:id2a knockdown Zebrafish retina with id2a morpholino treatment replicate 1 Filter out all reads with no call ‘N’ and low complexity reads without xxx 4 bases. Take reads that both paired end reads are passed the above condition. Map to EnsEMBL cDNA sequence then calculate RPK100M read counts per kb per 100 million of reads Summarize mapped reads per each cDNA. Determine differentially expressed genes with DESeq Genome build: EnsEMBL version 66 cDNA sequences Supplementary files format and content: tab delimited text files include RPKM values for each Sample DESeq output for differentially expressed genes. Zebrafish retina with id2a morpholino treatment replicate 1   Total RNA was isolated from dissected retinae at 48 hpf using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 microgram of total retinal RNA per condition to generate cDNA libraries by poly A selection.   tissue:retina|genotype/variation:id2a knockdown GSM949496 GSM949496: Zebrafish retina with id2a morpholino treatment replicate 1; Danio rerio; RNA Seq GSM949496 2 GSM949496: Zebrafish retina with id2a morpholino treatment replicate 1 1   GEO Accession:GSM949496 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>202</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC><READ_SPEC><READ_INDEX>1</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Reverse</READ_TYPE><BASE_COORD>102</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> SRP013815     Uribe201204_id2aKD1.untie_1.fastq.gz fastq 9529605934.0 47176267.0 GSM949496 r1 0:101 1:101 A:2628394362;C:2208991321;G:2220322548;T:2471897703;N:0 101 101     2628394362 2208991321 2220322548 2471897703 0 SRX155010 SRS346296 SRA054377 GEO Department of Biomedical Engineering, Ulsan National Institute of Science and Technology 2 0.83877 0.84035 0.0922 0.09318 0.76092 0.75795 0.48081 0.49029 101 101 B B biological fallback assumption illumina hiseq_era unknown poly_a trueseq bulk unknown unknown   South Korea 2012-06-18 Hatching Embryo Eye Sensory System
36383 36383 SRR514029 SRX155009 SRS346295 SRP013815 PRJNA168635 Id2a knockdown in zebrafish retina GSE38786 Transcriptome Analysis During vertebrate retinogenesis the precise balance between retinoblast proliferation and differentiation is spatially and temporally regulated through a number of intrinsic factors and extrinsic signaling pathways. Moreover there are complex gene regulatory network interactions between these intrinsic factors and extrinsic pathways which ultimately function to determine when retinoblasts exit the cell cycle and terminally differentiate. We recently uncovered a cell non autonomous role for the intrinsic HLH factor Id2a in regulating retinoblast proliferation and differentiation with Id2a deficient retinae containing an abundance of proliferative retinoblasts and an absence of terminally differentiated retinal neurons and glia. Here we report that Id2a function is necessary and sufficient to limit Notch pathway activity during retinogenesis. Id2a deficient retinae possess elevated levels of Notch pathway component gene expression while retinae overexpressing id2a possess reduced expression of Notch pathway component genes. Attenuation of Notch signaling activity by DAPT or by morpholino knockdown of Notch1a is sufficient to rescue both the proliferative and differentiation defects in Id2a deficient retinae. In addition to regulating Notch pathway activity through an RNA Seq and differential gene expression analysis of Id2a deficient retinae we identify a number of additional intrinsic and extrinsic regulatory pathway components whose expression is regulated by Id2a. These data highlight the integral role played by Id2a in the gene regulatory network governing the transition from retinoblast proliferation to terminal differentiation during vertebrate retinogenesis. Overall design: Two biological replicates for both Id2aMM and Id2aMO samples   pubmed:22981606   Zebrafish retina with Id2aMM morpholino treatment replicate 2 GSM949495   source name:Zebrafish retina with Id2aMM morpholino treatment replicate 2|tissue:retina|genotype/variation:control Zebrafish retina with Id2aMM morpholino treatment replicate 2 Filter out all reads with no call ‘N’ and low complexity reads without xxx 4 bases. Take reads that both paired end reads are passed the above condition. Map to EnsEMBL cDNA sequence then calculate RPK100M read counts per kb per 100 million of reads Summarize mapped reads per each cDNA. Determine differentially expressed genes with DESeq Genome build: EnsEMBL version 66 cDNA sequences Supplementary files format and content: tab delimited text files include RPKM values for each Sample DESeq output for differentially expressed genes. Zebrafish retina with Id2aMM morpholino treatment replicate 2   Total RNA was isolated from dissected retinae at 48 hpf using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 microgram of total retinal RNA per condition to generate cDNA libraries by poly A selection.   tissue:retina|genotype/variation:control GSM949495 GSM949495: Zebrafish retina with Id2aMM morpholino treatment replicate 2; Danio rerio; RNA Seq GSM949495 2 GSM949495: Zebrafish retina with Id2aMM morpholino treatment replicate 2 1   GEO Accession:GSM949495 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>202</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC><READ_SPEC><READ_INDEX>1</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Reverse</READ_TYPE><BASE_COORD>102</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> SRP013815     Uribe201204_ctrl2.untie_2.fastq.gz fastq 11102513678.0 54962939.0 GSM949495 r1 0:101 1:101 A:3114223438;C:2460129982;G:2471805247;T:3056355011;N:0 101 101     3114223438 2460129982 2471805247 3056355011 0 SRX155009 SRS346295 SRA054377 GEO Department of Biomedical Engineering, Ulsan National Institute of Science and Technology 2 0.86928 0.87394 0.14668 0.14966 0.73462 0.73428 0.47462 0.47615 101 101 B B biological fallback assumption illumina hiseq_era unknown poly_a trueseq bulk unknown unknown   South Korea 2012-06-18 Hatching Embryo Eye Sensory System
36384 36384 SRR514028 SRX155008 SRS346294 SRP013815 PRJNA168635 Id2a knockdown in zebrafish retina GSE38786 Transcriptome Analysis During vertebrate retinogenesis the precise balance between retinoblast proliferation and differentiation is spatially and temporally regulated through a number of intrinsic factors and extrinsic signaling pathways. Moreover there are complex gene regulatory network interactions between these intrinsic factors and extrinsic pathways which ultimately function to determine when retinoblasts exit the cell cycle and terminally differentiate. We recently uncovered a cell non autonomous role for the intrinsic HLH factor Id2a in regulating retinoblast proliferation and differentiation with Id2a deficient retinae containing an abundance of proliferative retinoblasts and an absence of terminally differentiated retinal neurons and glia. Here we report that Id2a function is necessary and sufficient to limit Notch pathway activity during retinogenesis. Id2a deficient retinae possess elevated levels of Notch pathway component gene expression while retinae overexpressing id2a possess reduced expression of Notch pathway component genes. Attenuation of Notch signaling activity by DAPT or by morpholino knockdown of Notch1a is sufficient to rescue both the proliferative and differentiation defects in Id2a deficient retinae. In addition to regulating Notch pathway activity through an RNA Seq and differential gene expression analysis of Id2a deficient retinae we identify a number of additional intrinsic and extrinsic regulatory pathway components whose expression is regulated by Id2a. These data highlight the integral role played by Id2a in the gene regulatory network governing the transition from retinoblast proliferation to terminal differentiation during vertebrate retinogenesis. Overall design: Two biological replicates for both Id2aMM and Id2aMO samples   pubmed:22981606   Zebrafish retina with Id2aMM morpholino treatment replicate 1 GSM949494   source name:Zebrafish retina with Id2aMM morpholino treatment replicate 1|tissue:retina|genotype/variation:control Zebrafish retina with Id2aMM morpholino treatment replicate 1 Filter out all reads with no call ‘N’ and low complexity reads without xxx 4 bases. Take reads that both paired end reads are passed the above condition. Map to EnsEMBL cDNA sequence then calculate RPK100M read counts per kb per 100 million of reads Summarize mapped reads per each cDNA. Determine differentially expressed genes with DESeq Genome build: EnsEMBL version 66 cDNA sequences Supplementary files format and content: tab delimited text files include RPKM values for each Sample DESeq output for differentially expressed genes. Zebrafish retina with Id2aMM morpholino treatment replicate 1   Total RNA was isolated from dissected retinae at 48 hpf using Trizol reagent. Illumina TruSeq RNA Sample Prep Kit was used with 1 microgram of total retinal RNA per condition to generate cDNA libraries by poly A selection.   tissue:retina|genotype/variation:control GSM949494 GSM949494: Zebrafish retina with Id2aMM morpholino treatment replicate 1; Danio rerio; RNA Seq GSM949494 2 GSM949494: Zebrafish retina with Id2aMM morpholino treatment replicate 1 1   GEO Accession:GSM949494 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 2000 <SPOT_DESCRIPTOR><SPOT_DECODE_SPEC><SPOT_LENGTH>202</SPOT_LENGTH><READ_SPEC><READ_INDEX>0</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Forward</READ_TYPE><BASE_COORD>1</BASE_COORD></READ_SPEC><READ_SPEC><READ_INDEX>1</READ_INDEX><READ_CLASS>Application Read</READ_CLASS><READ_TYPE>Reverse</READ_TYPE><BASE_COORD>102</BASE_COORD></READ_SPEC></SPOT_DECODE_SPEC></SPOT_DESCRIPTOR> SRP013815     Uribe201204_ctrl1.untie_1.fastq.gz fastq 9965568192.0 49334496.0 GSM949494 r1 0:101 1:101 A:2748582966;C:2261528563;G:2269950332;T:2685506331;N:0 101 101     2748582966 2261528563 2269950332 2685506331 0 SRX155008 SRS346294 SRA054377 GEO Department of Biomedical Engineering, Ulsan National Institute of Science and Technology 2 0.86051 0.86328 0.11721 0.11892 0.74184 0.74075 0.47603 0.47166 101 101 B B biological fallback assumption illumina hiseq_era unknown poly_a trueseq bulk unknown unknown   South Korea 2012-06-18 Hatching Embryo Eye Sensory System
49320 49320 SRR7886615 SRX4724267 SRS3809041 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60MGS3 GSM3397449   source name:60hpf GFAP:GFP Positive|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 60MGS3 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397449 GSM3397449: 60MGS3; Danio rerio; RNA Seq GSM3397449   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397449 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60MGS3.bam bam 2508986250.0 20071890.0 GSM3397449 r1 0:125 A:633150258;C:658892300;G:571953995;T:644986169;N:3528 125       633150258 658892300 571953995 644986169 3528 SRX4724267 SRS3809041 SRA780762 GEO University of Cambridge 1 0.0112   0.00292   0.98711   0.60601   125   T   under 1.2% mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49321 49321 SRR7886614 SRX4724266 SRS3809040 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60MGS2 GSM3397448   source name:60hpf GFAP:GFP Positive|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 60MGS2 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397448 GSM3397448: 60MGS2; Danio rerio; RNA Seq GSM3397448   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397448 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60MGS2.bam bam 2600602375.0 20804819.0 GSM3397448 r1 0:125 A:676529819;C:637723743;G:635912971;T:650421922;N:13920 125       676529819 637723743 635912971 650421922 13920 SRX4724266 SRS3809040 SRA780762 GEO University of Cambridge 1 0.01123   0.00282   0.98713   0.59463   125   T   under 1.2% mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49322 49322 SRR7886613 SRX4724265 SRS3809039 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60MGS1 GSM3397447   source name:60hpf GFAP:GFP Positive|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 60MGS1 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397447 GSM3397447: 60MGS1; Danio rerio; RNA Seq GSM3397447   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397447 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60MGS1.bam bam 2600602375.0 20804819.0 GSM3397447 r1 0:125 A:656042019;C:682911096;G:592791803;T:668797312;N:60145 125       656042019 682911096 592791803 668797312 60145 SRX4724265 SRS3809039 SRA780762 GEO University of Cambridge 1 0.01127   0.00304   0.98754   0.6105   125   T   under 1.2% mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49323 49323 SRR7886612 SRX4724264 SRS3809038 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60CS3 GSM3397446   source name:60hpf GFAP:GFP Negative|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 60CS3 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397446 GSM3397446: 60CS3; Danio rerio; RNA Seq GSM3397446   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397446 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60CS3.bam bam 3217257000.0 25738056.0 GSM3397446 r1 0:125 A:856598472;C:777828660;G:710658273;T:872152518;N:19077 125       856598472 777828660 710658273 872152518 19077 SRX4724264 SRS3809038 SRA780762 GEO University of Cambridge 1 0.4529   0.05353   0.79701   0.55977   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49324 49324 SRR7886611 SRX4724263 SRS3809037 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60CS2 GSM3397445   source name:60hpf GFAP:GFP Negative|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 60CS2 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397445 GSM3397445: 60CS2; Danio rerio; RNA Seq GSM3397445   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397445 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60CS2.bam bam 3308467000.0 26467736.0 GSM3397445 r1 0:125 A:898472326;C:766795565;G:762042071;T:881139599;N:17439 125       898472326 766795565 762042071 881139599 17439 SRX4724263 SRS3809037 SRA780762 GEO University of Cambridge 1 0.44529   0.05447   0.80273   0.53894   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49325 49325 SRR7886610 SRX4724262 SRS3809036 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   60CS1 GSM3397444   source name:60hpf GFAP:GFP Negative|age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 60CS1 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 60hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:60hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397444 GSM3397444: 60CS1; Danio rerio; RNA Seq GSM3397444   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397444 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 60CS1.bam bam 3308467000.0 26467736.0 GSM3397444 r1 0:125 A:881969824;C:799783674;G:727574215;T:899065817;N:73470 125       881969824 799783674 727574215 899065817 73470 SRX4724262 SRS3809036 SRA780762 GEO University of Cambridge 1 0.44178   0.05281   0.79849   0.54471   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49326 49326 SRR7886609 SRX4724261 SRS3809035 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48MGS3 GSM3397443   source name:48hpf GFAP:GFP Positive|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 48MGS3 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397443 GSM3397443: 48MGS3; Danio rerio; RNA Seq GSM3397443   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397443 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48MGS3.bam bam 3751483125.0 30011865.0 GSM3397443 r1 0:125 A:1063899440;C:813181012;G:740633593;T:1133592737;N:176343 125       1063899440 813181012 740633593 1133592737 176343 SRX4724261 SRS3809035 SRA780762 GEO University of Cambridge 1 0.87448   0.46322   0.85437   0.46617   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49327 49327 SRR7886608 SRX4724260 SRS3809034 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48MGS2 GSM3397442   source name:48hpf GFAP:GFP Positive|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 48MGS2 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397442 GSM3397442: 48MGS2; Danio rerio; RNA Seq GSM3397442   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397442 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48MGS2.bam bam 3253082750.0 26024662.0 GSM3397442 r1 0:125 A:860727696;C:787565434;G:719641544;T:885087745;N:60331 125       860727696 787565434 719641544 885087745 60331 SRX4724260 SRS3809034 SRA780762 GEO University of Cambridge 1 0.70949   0.06764   0.79251   0.56599   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49328 49328 SRR7886607 SRX4724259 SRS3809033 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48MGS1 GSM3397441   source name:48hpf GFAP:GFP Positive|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive 48MGS1 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Positive   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Positive GSM3397441 GSM3397441: 48MGS1; Danio rerio; RNA Seq GSM3397441   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397441 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48MGS1.bam bam 3155766875.0 25246135.0 GSM3397441 r1 0:125 A:835678187;C:762799231;G:696339226;T:860875540;N:74691 125       835678187 762799231 696339226 860875540 74691 SRX4724259 SRS3809033 SRA780762 GEO University of Cambridge 1 0.70755   0.06855   0.79569   0.53396   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49329 49329 SRR7886606 SRX4724258 SRS3809032 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48CS3 GSM3397440   source name:48hpf GFAP:GFP Negative|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 48CS3 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397440 GSM3397440: 48CS3; Danio rerio; RNA Seq GSM3397440   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397440 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48CS3.bam bam 662621625.0 5300973.0 GSM3397440 r1 0:125 A:179417199;C:153458866;G:152649509;T:177095017;N:1034 125       179417199 153458866 152649509 177095017 1034 SRX4724258 SRS3809032 SRA780762 GEO University of Cambridge 1 0.70566   0.06917   0.79488   0.58099   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49330 49330 SRR7886605 SRX4724257 SRS3809031 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48CS2 GSM3397439   source name:48hpf GFAP:GFP Negative|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 48CS2 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397439 GSM3397439: 48CS2; Danio rerio; RNA Seq GSM3397439   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397439 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48CS2.bam bam 959630000.0 7677040.0 GSM3397439 r1 0:125 A:250545467;C:236640013;G:213639661;T:258800686;N:4173 125       250545467 236640013 213639661 258800686 4173 SRX4724257 SRS3809031 SRA780762 GEO University of Cambridge 1 0.55605   0.07819   0.80186   0.59736   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
49331 49331 SRR7886604 SRX4724256 SRS3809030 SRP162288 PRJNA492363 Genetic control of cellular morphogenesis in Müller glia GSE120275 Transcriptome Analysis How the various cell types of the body achieve their specific shapes is fundamentally unknown. Here we explore this issue by identifying genes involved in the elaboration of the complex yet conserved cellular morphology of Müller glial MG cells in the retina. Using genomic based strategies in zebrafish we found more than 40 candidate genes involved in specific aspects of MG morphogenesis. The successive steps of cell morphogenesis correlate with the timing of the expression of cohorts of inter related genes that have roles in generating the particular anatomical features of these cells suggesting that a sequence of genetic regulomes govern stepwise cellular morphogenesis in this system. Overall design: 12 samples with three replicates each are provided. GFAP:GFP positive and negative cells were FAC sorted from wild type animals from each developmental stage   pubmed:30924555   48CS1 GSM3397438   source name:48hpf GFAP:GFP Negative|age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative 48CS1 Trimmomatic read trimming Hisat2 genome aligment Samtools Sam to Bam sorting Bioconuducter Feature counts Rsubread limma DESeq2 DEFormats org.Dr.eg.db EdgeR Genome build: GRCz10 Supplementary files format and content: TMM fold change analysis 48hpf GFAP:GFP Negative   Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer’s protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions   age:48hpf|genotype:Control GFAP:GFP|tissue:Dissociated retinal tissue|gfap:Negative GSM3397438 GSM3397438: 48CS1; Danio rerio; RNA Seq GSM3397438   1 Single cell suspensions were sorted on a Beckman Coulter MoFlo to capture Muller glia GFP and control retinal tissue non GFP. Cells were sorted into lysis buffer and RNA was immediately extracted using the RNeasy mini kit Qiagen. RNA concentration and qualities were assessed on an Agilent Bioanalyzer and RNA amplification and cDNA synthesis was performed with the Ovation RNA Amplification System V2 NuGEN using manufacturer's protocol. Nextera library preparations were performed using the Nextera DNA library kit according to the manufacturer's directions GEO Accession:GSM3397438 RNA-Seq TRANSCRIPTOMIC cDNA SINGLE ILLUMINA Illumina HiSeq 2500   SRP162288   dangling references:treat as unmapped 48CS1.bam bam 959630000.0 7677040.0 GSM3397438 r1 0:125 A:248562218;C:241973573;G:206301549;T:262772093;N:20567 125       248562218 241973573 206301549 262772093 20567 SRX4724256 SRS3809030 SRA780762 GEO University of Cambridge 1 0.55246   0.07651   0.79659   0.6137   125   B   usable mapping rate illumina hiseq_era unknown cdna_unspecified nextera bulk unknown unknown   United Kingdom 2018-09-20 Hatching Embryo Eye Sensory System
50656 50656 SRR10769144 SRX7443062 SRS5887360 SRP169565 PRJNA506002 Unifying Developmental Programs for Embryonic and Post Embryonic Neurogenesis in the Zebrafish Retina GSE122680 Transcriptome Analysis It provides an opportunity to make the first comparative study of molecular programs used by embryonic and post embryonic neurogenic programs in the vertebrate retina. Overall design: Retinal samples from 4 developmental stages and 1 post embryonic stage   pubmed:32467236   48 hpf retina rep GSM4233148   source name:Retina|strain:AB|genotype:wildtype|tissue:whole retina|developmental stage:48 hpf 48 hpf retina rep Single cell FASTQ sequencing reads Novogene aligned to the zebrafish genome Zv10 were processed and converted to digital gene expression matrices using the Cell Ranger Single Cell Software Suite v2.1.0 provided in 10x genomics website https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger. To further analyze the data we used an analysis pipeline provided by the Seurat R package http://satijalab.org/seurat/. We subtract progenitor cell clusters of 36hpf and 48hpf samples according to the proliferative gene expression. For post embryonic samples we subtract the ciliary marginal zone CMZ cell clusters based on marker genes' expression. Genome build: Zv10 Supplementary files format and content: barcodes.tsv genes.tsv matrix.mtx Retina   We dissected 24 36 48 72 hpf retinas of wild type zebrafishes and isolated via papin solution. 14 dpf retinas were dissected from PCNA GFP transgenic fishes. And post isolation by papin solution the GFP positive cells which could label the ciliary marginal zone CMZ cells were sorted via FACS. These isolated cells filtered by 40μm cell strainer BD Falcon were loaded onto the Chromium Single Cell Chip10x Genomics according to the manufacturer’s protocol. Single cell RNA seq libraries were generated using the GemCode Single Cell Instrument and Single Cell three prime Library &Gel Bead kit v2 Chip kit 10x Genomics following the manufacturer’s protocol.   strain:AB|genotype:wildtype|tissue:whole retina|developmental stage:48 hpf GSM4233148 GSM4233148: 48 hpf retina rep; Danio rerio; RNA Seq GSM4233148   1 We dissected 24 36 48 72 hpf retinas of wild type zebrafishes and isolated via papin solution. 14 dpf retinas were dissected from PCNA GFP transgenic fishes. And post isolation by papin solution the GFP positive cells which could label the ciliary marginal zone CMZ cells were sorted via FACS. These isolated cells filtered by 40μm cell strainer BD Falcon were loaded onto the Chromium Single Cell Chip10x Genomics according to the manufacturer's protocol. Single cell RNA seq libraries were generated using the GemCode Single Cell Instrument and Single Cell three prime Library &Gel Bead kit v2 Chip kit 10x Genomics following the manufacturer's protocol. GEO Accession:GSM4233148 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP169565   intentional duplicate 48hpf_rep_possorted_genome_bam.bam 10X Genomics bam file 31477860150.0 209852401.0 GSM4233148 r1 0:150 A:9531250692;C:6384514147;G:7053602060;T:8508157890;N:335361 150       9531250692 6384514147 7053602060 8508157890 335361 SRX7443062 SRS5887360 SRA812747 GEO Lab of Stem Cell and Neurogenesis, Institute of Neuroscience, CAS 1 0.91107   0.07208   0.83644   0.47681   150   B   usable mapping rate illumina novaseq_era unknown cdna_unspecified unknown sc single_cell_droplet 10x   China 2019-12-26 Hatching Embryo Eye Sensory System
50661 50661 SRR11812054 SRX5025779 SRS4057279 SRP169565 PRJNA506002 Unifying Developmental Programs for Embryonic and Post Embryonic Neurogenesis in the Zebrafish Retina GSE122680 Transcriptome Analysis It provides an opportunity to make the first comparative study of molecular programs used by embryonic and post embryonic neurogenic programs in the vertebrate retina. Overall design: Retinal samples from 4 developmental stages and 1 post embryonic stage   pubmed:32467236   48 hpf retina GSM3478015   source name:Retina|strain:AB|genotype:wildtype|tissue:whole retina|developmental stage:48 hpf 48 hpf retina Single cell FASTQ sequencing reads Novogene aligned to the zebrafish genome Zv10 were processed and converted to digital gene expression matrices using the Cell Ranger Single Cell Software Suite v2.1.0 provided in 10x genomics website https://support.10xgenomics.com/single cell gene expression/software/pipelines/latest/what is cell ranger. To further analyze the data we used an analysis pipeline provided by the Seurat R package http://satijalab.org/seurat/. We substract progenitor cell clusters of 36hpf and 48hpf samples according to the proliferative gene expression. For post embryonic samples we substract the ciliary marginal zone CMZ cell clusters based on marker genes' expression. Genome build: Zv10 Supplementary files format and content: barcodes.tsv genes.tsv matrix.mtx Retina   We dissected 24 36 48 72 hpf retinas of wild type zebrafishes and isolated via papin solution. 14 dpf retinas were dissected from PCNA GFP transgenic fishes. And post isolation by papin solution the GFP positive cells which could label the ciliary marginal zone CMZ cells were sorted via FACS. These isolated cells filtered by 40μm cell strainer BD Falcon were loaded onto the Chromium Single Cell Chip10x Genomics according to the manufacturer’s protocol. Single cell RNA seq libraries were generated using the GemCode Single Cell Instrument and Single Cell three prime Library &Gel Bead kit v2 Chip kit 10x Genomics following the manufacturer’s protocol.   strain:AB|genotype:wildtype|tissue:whole retina|developmental stage:48 hpf GSM3478015 GSM3478015: 48 hpf retina; Danio rerio; RNA Seq GSM3478015   1 We dissected 24 36 48 72 hpf retinas of wild type zebrafishes and isolated via papin solution. 14 dpf retinas were dissected from PCNA GFP transgenic fishes. And post isolation by papin solution the GFP positive cells which could label the ciliary marginal zone CMZ cells were sorted via FACS. These isolated cells filtered by 40μm cell strainer BD Falcon were loaded onto the Chromium Single Cell Chip10x Genomics according to the manufacturer's protocol. Single cell RNA seq libraries were generated using the GemCode Single Cell Instrument and Single Cell three prime Library &Gel Bead kit v2 Chip kit 10x Genomics following the manufacturer's protocol. GEO Accession:GSM3478015 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina NovaSeq 6000   SRP169565   intentional duplicate 48hpf_possorted_genome_bam.bam 10X Genomics bam file 27828827400.0 185525516.0 GSM3478015 r11 0:150 A:8568529313;C:5431445717;G:5968188905;T:7860573445;N:90020 150       8568529313 5431445717 5968188905 7860573445 90020 SRX5025779 SRS4057279 SRA812747 GEO Lab of Stem Cell and Neurogenesis, Institute of Neuroscience, CAS 1 0.91308   0.11004   0.82213   0.48772   150   B   usable mapping rate illumina novaseq_era unknown cdna_unspecified unknown sc single_cell_droplet 10x   China 2018-11-19 Hatching Embryo Eye Sensory System
61581 61581 SRR12874662 SRX9340686 SRS7562493 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 56 hpf Rep5 [5 56 R] GSM4848085   source name:dorsal retina 56 hpf|hpf:56|tissue:dorsal retina DR 56 hpf Rep5 [5 56 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:dorsal retina GSM4848085 GSM4848085: DR 56 hpf Rep5 [5 56 R]; Danio rerio; RNA Seq GSM4848085   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848085 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     5-56-R_R1.fastq.gz 5-56-R_R2.fastq.gz fastq fastq 3021678200.0 15108391.0 GSM4848085 r1 0:100 1:100 A:842653749;C:670303060;G:690387728;T:815456479;N:2877184 100 100     842653749 670303060 690387728 815456479 2877184 SRX9340686 SRS7562493 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94619 0.94623 0.15083 0.1507 0.74639 0.74834 0.48701 0.49558 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61582 61582 SRR12874661 SRX9340685 SRS7562495 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 56 hpf Rep5 [5 56 0] GSM4848084   source name:optic fissure retina 56 hpf|hpf:56|tissue:optic fissure retina OF 56 hpf Rep5 [5 56 0] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:optic fissure retina GSM4848084 GSM4848084: OF 56 hpf Rep5 [5 56 0]; Danio rerio; RNA Seq GSM4848084   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848084 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     5-56-O_R1.fastq.gz 5-56-O_R2.fastq.gz fastq fastq 2975878600.0 14879393.0 GSM4848084 r1 0:100 1:100 A:837964142;C:651309804;G:674797335;T:808675013;N:3132306 100 100     837964142 651309804 674797335 808675013 3132306 SRX9340685 SRS7562495 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94334 0.94301 0.17167 0.17156 0.7376 0.73764 0.48724 0.48286 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61583 61583 SRR12874660 SRX9340684 SRS7562494 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 48 hpf Rep5 [5 48 R] GSM4848083   source name:dorsal retina 48 hpf|hpf:48|tissue:dorsal retina DR 48 hpf Rep5 [5 48 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:dorsal retina GSM4848083 GSM4848083: DR 48 hpf Rep5 [5 48 R]; Danio rerio; RNA Seq GSM4848083   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848083 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     5-48-R_R1.fastq.gz 5-48-R_R2.fastq.gz fastq fastq 2239921200.0 11199606.0 GSM4848083 r1 0:100 1:100 A:625661378;C:494955850;G:511675170;T:605386182;N:2242620 100 100     625661378 494955850 511675170 605386182 2242620 SRX9340684 SRS7562494 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94552 0.9457 0.15214 0.1519 0.7431 0.74381 0.49047 0.48932 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61584 61584 SRR12874659 SRX9340683 SRS7562492 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 48 hpf Rep5 [5 48 O] GSM4848082   source name:optic fissure retina 48 hpf|hpf:48|tissue:optic fissure retina OF 48 hpf Rep5 [5 48 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:optic fissure retina GSM4848082 GSM4848082: OF 48 hpf Rep5 [5 48 O]; Danio rerio; RNA Seq GSM4848082   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848082 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     5-48-O_R1.fastq.gz 5-48-O_R2.fastq.gz fastq fastq 2833605600.0 14168028.0 GSM4848082 r1 0:100 1:100 A:788130966;C:630887942;G:649916615;T:761868221;N:2801856 100 100     788130966 630887942 649916615 761868221 2801856 SRX9340683 SRS7562492 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94695 0.94729 0.14775 0.14824 0.73953 0.74164 0.49175 0.4918 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61587 61587 SRR12874656 SRX9340680 SRS7562490 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 56 hpf Rep4 [4 56 R] GSM4848079   source name:dorsal retina 56 hpf|hpf:56|tissue:dorsal retina DR 56 hpf Rep4 [4 56 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:dorsal retina GSM4848079 GSM4848079: DR 56 hpf Rep4 [4 56 R]; Danio rerio; RNA Seq GSM4848079   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848079 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     4-56-R_R1.fastq.gz 4-56-R_R2.fastq.gz fastq fastq 2575355000.0 12876775.0 GSM4848079 r1 0:100 1:100 A:719372734;C:569208001;G:591353722;T:692825451;N:2595092 100 100     719372734 569208001 591353722 692825451 2595092 SRX9340680 SRS7562490 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94196 0.94214 0.14248 0.14239 0.74519 0.74556 0.48766 0.48749 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61588 61588 SRR12874655 SRX9340679 SRS7562488 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 56 hpf Rep4 [4 56 O] GSM4848078   source name:optic fissure retina 56 hpf|hpf:56|tissue:optic fissure retina OF 56 hpf Rep4 [4 56 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:optic fissure retina GSM4848078 GSM4848078: OF 56 hpf Rep4 [4 56 O]; Danio rerio; RNA Seq GSM4848078   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848078 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     4-56-O_R1.fastq.gz 4-56-O_R2.fastq.gz fastq fastq 2962950600.0 14814753.0 GSM4848078 r1 0:100 1:100 A:839363055;C:643262435;G:667200849;T:810187958;N:2936303 100 100     839363055 643262435 667200849 810187958 2936303 SRX9340679 SRS7562488 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93382 0.93302 0.17575 0.17531 0.74085 0.74223 0.49824 0.504 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61589 61589 SRR12874654 SRX9340678 SRS7562487 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 48 hpf Rep4 [4 48 R] GSM4848077   source name:dorsal retina 48 hpf|hpf:48|tissue:dorsal retina DR 48 hpf Rep4 [4 48 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:dorsal retina GSM4848077 GSM4848077: DR 48 hpf Rep4 [4 48 R]; Danio rerio; RNA Seq GSM4848077   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848077 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     4-48-R_R1.fastq.gz 4-48-R_R2.fastq.gz fastq fastq 2844671200.0 14223356.0 GSM4848077 r1 0:100 1:100 A:794974141;C:628655166;G:651606298;T:766506062;N:2929533 100 100     794974141 628655166 651606298 766506062 2929533 SRX9340678 SRS7562487 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94228 0.94286 0.14952 0.1508 0.7558 0.75621 0.49161 0.49112 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61590 61590 SRR12874653 SRX9340677 SRS7562486 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 48 hpf Rep4 [4 48 O] GSM4848076   source name:optic fissure retina 48 hpf|hpf:48|tissue:optic fissure retina OF 48 hpf Rep4 [4 48 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:optic fissure retina GSM4848076 GSM4848076: OF 48 hpf Rep4 [4 48 O]; Danio rerio; RNA Seq GSM4848076   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848076 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     4-48-O_R1.fastq.gz 4-48-O_R2.fastq.gz fastq fastq 2972787000.0 14863935.0 GSM4848076 r1 0:100 1:100 A:831928443;C:656429034;G:674981859;T:806440139;N:3007525 100 100     831928443 656429034 674981859 806440139 3007525 SRX9340677 SRS7562486 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94047 0.93996 0.16117 0.1608 0.743 0.74533 0.48986 0.49728 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61593 61593 SRR12874650 SRX9340674 SRS7562482 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 56 hpf Rep3 [3 56 R] GSM4848073   source name:dorsal retina 56 hpf|hpf:56|tissue:dorsal retina DR 56 hpf Rep3 [3 56 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:dorsal retina GSM4848073 GSM4848073: DR 56 hpf Rep3 [3 56 R]; Danio rerio; RNA Seq GSM4848073   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848073 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     3-56-R_R1.fastq.gz 3-56-R_R2.fastq.gz fastq fastq 2836655800.0 14183279.0 GSM4848073 r1 0:100 1:100 A:796999987;C:622654837;G:648405815;T:765641148;N:2954013 100 100     796999987 622654837 648405815 765641148 2954013 SRX9340674 SRS7562482 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94067 0.94077 0.15638 0.15562 0.74403 0.74497 0.49593 0.49546 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61594 61594 SRR12874649 SRX9340673 SRS7562484 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 56 hpf Rep3 [3 56 O] GSM4848072   source name:optic fissure retina 56 hpf|hpf:56|tissue:optic fissure retina OF 56 hpf Rep3 [3 56 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:optic fissure retina GSM4848072 GSM4848072: OF 56 hpf Rep3 [3 56 O]; Danio rerio; RNA Seq GSM4848072   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848072 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     3-56-O_R1.fastq.gz 3-56-O_R2.fastq.gz fastq fastq 2962222200.0 14811111.0 GSM4848072 r1 0:100 1:100 A:839522002;C:642451913;G:665094462;T:812146216;N:3007607 100 100     839522002 642451913 665094462 812146216 3007607 SRX9340673 SRS7562484 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93685 0.93543 0.17034 0.17106 0.73914 0.74087 0.49394 0.4949 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61595 61595 SRR12874648 SRX9340672 SRS7562481 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 48 hpf Rep3 [3 48 R] GSM4848071   source name:dorsal retina 48 hpf|hpf:48|tissue:dorsal retina DR 48 hpf Rep3 [3 48 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:dorsal retina GSM4848071 GSM4848071: DR 48 hpf Rep3 [3 48 R]; Danio rerio; RNA Seq GSM4848071   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848071 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     3-48-R_R1.fastq.gz 3-48-R_R2.fastq.gz fastq fastq 3015560600.0 15077803.0 GSM4848071 r1 0:100 1:100 A:857335603;C:650996829;G:676858466;T:827306581;N:3063121 100 100     857335603 650996829 676858466 827306581 3063121 SRX9340672 SRS7562481 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.92909 0.92812 0.17933 0.17852 0.75708 0.75903 0.45933 0.50484 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61596 61596 SRR12874647 SRX9340671 SRS7562480 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 48 hpf Rep3 [3 48 O] GSM4848070   source name:optic fissure retina 48 hpf|hpf:48|tissue:optic fissure retina OF 48 hpf Rep3 [3 48 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:optic fissure retina GSM4848070 GSM4848070: OF 48 hpf Rep3 [3 48 O]; Danio rerio; RNA Seq GSM4848070   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848070 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     3-48-O_R1.fastq.gz 3-48-O_R2.fastq.gz fastq fastq 3617936400.0 18089682.0 GSM4848070 r1 0:100 1:100 A:1017979863;C:791826181;G:821834684;T:982508058;N:3787614 100 100     1017979863 791826181 821834684 982508058 3787614 SRX9340671 SRS7562480 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93482 0.93556 0.17727 0.1779 0.73217 0.73267 0.49331 0.49384 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61599 61599 SRR12874644 SRX9340668 SRS7562477 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 56 hpf Rep2 [2 56 R] GSM4848067   source name:dorsal retina 56 hpf|hpf:56|tissue:dorsal retina DR 56 hpf Rep2 [2 56 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:dorsal retina GSM4848067 GSM4848067: DR 56 hpf Rep2 [2 56 R]; Danio rerio; RNA Seq GSM4848067   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848067 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     2-56-R_R1.fastq.gz 2-56-R_R2.fastq.gz fastq fastq 2736567000.0 13682835.0 GSM4848067 r1 0:100 1:100 A:774624618;C:594658601;G:615415962;T:749131027;N:2736792 100 100     774624618 594658601 615415962 749131027 2736792 SRX9340668 SRS7562477 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93368 0.93451 0.17403 0.17569 0.74201 0.74479 0.5181 0.50551 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61600 61600 SRR12874643 SRX9340667 SRS7562476 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 56 hpf Rep2 [2 56 O] GSM4848066   source name:optic fissure retina 56 hpf|hpf:56|tissue:optic fissure retina OF 56 hpf Rep2 [2 56 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:optic fissure retina GSM4848066 GSM4848066: OF 56 hpf Rep2 [2 56 O]; Danio rerio; RNA Seq GSM4848066   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848066 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     2-56-O_R1.fastq.gz 2-56-O_R2.fastq.gz fastq fastq 2676334400.0 13381672.0 GSM4848066 r1 0:100 1:100 A:755448065;C:583050210;G:604376373;T:730670069;N:2789683 100 100     755448065 583050210 604376373 730670069 2789683 SRX9340667 SRS7562476 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93546 0.93514 0.19101 0.19201 0.72088 0.7217 0.4938 0.48928 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61601 61601 SRR12874642 SRX9340666 SRS7562475 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 48 hpf Rep2 [2 48 R] GSM4848065   source name:dorsal retina 48 hpf|hpf:48|tissue:dorsal retina DR 48 hpf Rep2 [2 48 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:dorsal retina GSM4848065 GSM4848065: DR 48 hpf Rep2 [2 48 R]; Danio rerio; RNA Seq GSM4848065   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848065 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     2-48-R_R1.fastq.gz 2-48-R_R2.fastq.gz fastq fastq 3092140600.0 15460703.0 GSM4848065 r1 0:100 1:100 A:865594525;C:682549898;G:702370234;T:838685013;N:2940930 100 100     865594525 682549898 702370234 838685013 2940930 SRX9340666 SRS7562475 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.94438 0.94383 0.14587 0.1461 0.74211 0.74464 0.50115 0.50098 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61602 61602 SRR12874641 SRX9340665 SRS7562474 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 48 hpf Rep2 [2 48 O] GSM4848064   source name:optic fissure retina 48 hpf|hpf:48|tissue:optic fissure retina OF 48 hpf Rep2 [2 48 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:optic fissure retina GSM4848064 GSM4848064: OF 48 hpf Rep2 [2 48 O]; Danio rerio; RNA Seq GSM4848064   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848064 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     2-48-O_R1.fastq.gz 2-48-O_R2.fastq.gz fastq fastq 2606582200.0 13032911.0 GSM4848064 r1 0:100 1:100 A:739206672;C:564781083;G:586126893;T:713698007;N:2769545 100 100     739206672 564781083 586126893 713698007 2769545 SRX9340665 SRS7562474 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93607 0.93608 0.18238 0.18202 0.72719 0.72827 0.49891 0.49686 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61605 61605 SRR12874638 SRX9340662 SRS7562471 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 56 hpf Rep1 [1 56 R] GSM4848061   source name:dorsal retina 56 hpf|hpf:56|tissue:dorsal retina DR 56 hpf Rep1 [1 56 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:dorsal retina GSM4848061 GSM4848061: DR 56 hpf Rep1 [1 56 R]; Danio rerio; RNA Seq GSM4848061   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848061 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     1-56-R_R1.fastq.gz 1-56-R_R2.fastq.gz fastq fastq 2657160200.0 13285801.0 GSM4848061 r1 0:100 1:100 A:783255245;C:545277836;G:561089398;T:764970589;N:2567132 100 100     783255245 545277836 561089398 764970589 2567132 SRX9340662 SRS7562471 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.89392 0.89321 0.38815 0.38896 0.70516 0.70565 0.5057 0.49386 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61606 61606 SRR12874637 SRX9340661 SRS7562470 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 56 hpf Rep1 [1 56 O] GSM4848060   source name:optic fissure retina 56 hpf|hpf:56|tissue:optic fissure retina OF 56 hpf Rep1 [1 56 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 56 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:56|tissue:optic fissure retina GSM4848060 GSM4848060: OF 56 hpf Rep1 [1 56 O]; Danio rerio; RNA Seq GSM4848060   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848060 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     1-56-O_R1.fastq.gz 1-56-O_R2.fastq.gz fastq fastq 2610698400.0 13053492.0 GSM4848060 r1 0:100 1:100 A:747164488;C:558793373;G:578023357;T:723973829;N:2743353 100 100     747164488 558793373 578023357 723973829 2743353 SRX9340661 SRS7562470 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.92606 0.92637 0.20702 0.20634 0.72056 0.72121 0.50668 0.50793 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61607 61607 SRR12874636 SRX9340660 SRS7562469 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   DR 48 hpf Rep1 [1 48 R] GSM4848059   source name:dorsal retina 48 hpf|hpf:48|tissue:dorsal retina DR 48 hpf Rep1 [1 48 R] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample dorsal retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:dorsal retina GSM4848059 GSM4848059: DR 48 hpf Rep1 [1 48 R]; Danio rerio; RNA Seq GSM4848059   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848059 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     1-48-R_R1.fastq.gz 1-48-R_R2.fastq.gz fastq fastq 2342612000.0 11713060.0 GSM4848059 r1 0:100 1:100 A:660019342;C:511509124;G:533952026;T:634790133;N:2341375 100 100     660019342 511509124 533952026 634790133 2341375 SRX9340660 SRS7562469 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93651 0.93755 0.17081 0.17042 0.74194 0.74322 0.50319 0.50384 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
61608 61608 SRR12874635 SRX9340659 SRS7562467 SRP288083 PRJNA670550 Transcriptome profiling of zebrafish optic fissure fusion GSE159822 Transcriptome Analysis Incomplete fusion of the optic fissure leads to ocular coloboma a congenital eye defect that affects up to 7.5 per 10 000 births and accounts for up to 10 percent of childhood blindness. The molecular and cellular mechanisms that facilitate optic fissure fusion remain elusive. We have profiled global gene expression during optic fissure morphogenesis by transcriptome analysis of tissue dissected from the margins of the zebrafish optic fissure and the opposing dorsal retina before 32 hpf hpf during 48 hpf and post 56 hpf optic fissure fusion. Differential expression analysis between optic fissure and dorsal retinal tissue resulted in the detection of several known and novel developmental genes. The expression of selected genes was validated by qRT PCR analysis and localisation investigated using in situ hybridisation. We discuss significantly overrepresented functional ontology categories in the context of optic fissure morphogenesis and highlight interesting transcripts from hierarchical clustering for subsequent analysis. We have identified netrin1a ntn1a as highly differentially expressed across optic fissure fusion with a resultant ocular coloboma phenotype following morpholino antisense translation blocking knockdown and downstream disruption of atoh7 expression. To support the identification of candidate genes in human studies we have generated an online open access resource for fast and simple quantitative querying of the gene expression data. Our study represents the first comprehensive analysis of the zebrafish optic fissure transcriptome and provides a valuable resource to facilitate our understanding of the complex aetiology of ocular coloboma. Overall design: We used RNA seq to identify differentially expressed genes across optic fissure fusion in zebrafish. Samples were isolated retinal regions including the optic fissure OF region and control dorsal retina DR at timepoints; prefusion 32 hpf fusing 48 hpf and post fusion 56 hpf. 5 biological replicates.   pubmed:30733552;pubmed:35034853   OF 48 hpf Rep1 [1 48 O] GSM4848058   source name:optic fissure retina 48 hpf|hpf:48|tissue:optic fissure retina OF 48 hpf Rep1 [1 48 O] Illumina Casava1.7 software used for basecalling. Sequenced reads were trimmed for adaptor sequence and masked for low complexity or low quality sequence using trimgalore Reads were aligned to the genome and transcriptome using STAR v2.4.0 using parameters outSAMtype BAM Unsorted outSAMunmapped Within Resulting BAM files were sorted by coodinates and duplicates soft marked with picard MarkDuplicates Count files were created using htseq count v0.6.1 and modelled using DESeq2 Genome build: GRCz10 Supplementary files format and content: tab delimited text files include raw count values for each sample optic fissure retina 48 hpf   RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols zebrafish were harvested at appropriate time points hpf retinal regions dissected hpf:48|tissue:optic fissure retina GSM4848058 GSM4848058: OF 48 hpf Rep1 [1 48 O]; Danio rerio; RNA Seq GSM4848058   1 RNA was extracted using Qiagen Rneasy FFPE kit and quantified RNA seq libraries were constructed using the SMARTer low input kit using standard protocols GEO Accession:GSM4848058 RNA-Seq TRANSCRIPTOMIC cDNA PAIRED ILLUMINA Illumina HiSeq 4000   SRP288083     1-48-O_R1.fastq.gz 1-48-O_R2.fastq.gz fastq fastq 2552005400.0 12760027.0 GSM4848058 r1 0:100 1:100 A:720990425;C:556371737;G:577499015;T:694589372;N:2554851 100 100     720990425 556371737 577499015 694589372 2554851 SRX9340659 SRS7562467 SRA1146222 GEO Institute of Ophthalmology, UCL 2 0.93333 0.93374 0.1828 0.18383 0.73708 0.73841 0.49612 0.49916 100 100 B B biological fallback assumption illumina hiseq_era full_length cdna_unspecified smarter bulk unknown unknown   United Kingdom 2020-10-21 Hatching Embryo Eye Sensory System
69475 69475 SRR18712324 SRX14812927 SRS12569570 SRP369458 PRJNA825495 Regulation of retina microexons by srrm3 in zebrafish PRJNA825495 Other To investigate the role of srrm3 as a regulator of retina microexons RetMICs we have sequenced heads from WT animals at 24 hpf and enucleated eyes from 48 hpf 60 hpf and 72 hpf larvae WT and KO for srrm3.         Eye 60hpf WT b   strain:WT|dev stage:60 hpf|sex:NA|tissue:Eye|BioSampleModel:Model organism or animal                 Eyes from 60 hpf larvae WT for the eMIC domain of srrm3. Eye 60hpf WT Eye 60hpf WT WT and srrm3 mutant zebrafish were grown at 28C 14h light/10h dark cycle. Extract protocol: 24 hpf WT fish heads were removed and pulled for RNA extraction using RNA RNeasy Mini kit QIAGEN. Larvae eyes were removed and from twenty to sixty eyes per genotype were pulled for RNA extraction using RNA RNeasy Micro kit QIAGENLibrary preparation: RNA libraries were prepared for sequencing using standard Illumina protocols; polyA selected stranded RNA seq     RNA-Seq TRANSCRIPTOMIC RANDOM PAIRED ILLUMINA Illumina HiSeq 2500   SRP369458     Eye_60hpf_WT_b_R1-125.fq.gz Eye_60hpf_WT_b_R2-125.fq.gz fastq fastq 15250592500.0 61002370.0 Eye 60hpf WT b R1 125.fq.gz 0:125 1:125 A:3923837289;C:3702942025;G:3742033933;T:3872922356;N:8856897 125 125     3923837289 3702942025 3742033933 3872922356 8856897 SRX14812927 SRS12569570 SRA1402039 Centre for Genomic Regulation|Systems Biology Centre for Genomic Regulation|Systems Biology 2 0.97168 0.9731 0.0527 0.05258 0.69891 0.70041 0.45973 0.45381 125 125 B B biological fallback assumption illumina hiseq_era unknown random_priming trueseq bulk unknown unknown   Spain 2022-04-11 Hatching Embryo Eye Sensory System
69476 69476 SRR18712325 SRX14812926 SRS12569569 SRP369458 PRJNA825495 Regulation of retina microexons by srrm3 in zebrafish PRJNA825495 Other To investigate the role of srrm3 as a regulator of retina microexons RetMICs we have sequenced heads from WT animals at 24 hpf and enucleated eyes from 48 hpf 60 hpf and 72 hpf larvae WT and KO for srrm3.         Eye 60hpf MUT b   strain:srrm3 MUT|dev stage:60 hpf|sex:NA|tissue:Eye|BioSampleModel:Model organism or animal                 Eyes from 60 hpf larvae mutated for the eMIC domain of srrm3. Eye 60hpf MUT Eye 60hpf MUT WT and srrm3 mutant zebrafish were grown at 28C 14h light/10h dark cycle. Extract protocol: 24 hpf WT fish heads were removed and pulled for RNA extraction using RNA RNeasy Mini kit QIAGEN. Larvae eyes were removed and from twenty to sixty eyes per genotype were pulled for RNA extraction using RNA RNeasy Micro kit QIAGENLibrary preparation: RNA libraries were prepared for sequencing using standard Illumina protocols; polyA selected stranded RNA seq     RNA-Seq TRANSCRIPTOMIC RANDOM PAIRED ILLUMINA Illumina HiSeq 2500   SRP369458     Eye_60hpf_MUT_b_R1-125.fq.gz Eye_60hpf_MUT_b_R2-125.fq.gz fastq fastq 14932544250.0 59730177.0 Eye 60hpf MUT b R1 125.fq.gz 0:125 1:125 A:3891825609;C:3575114135;G:3598083383;T:3858808039;N:8713084 125 125     3891825609 3575114135 3598083383 3858808039 8713084 SRX14812926 SRS12569569 SRA1402039 Centre for Genomic Regulation|Systems Biology Centre for Genomic Regulation|Systems Biology 2 0.96985 0.97098 0.06006 0.0593 0.68939 0.6926 0.46001 0.45867 125 125 B B biological fallback assumption illumina hiseq_era unknown random_priming trueseq bulk unknown unknown   Spain 2022-04-11 Hatching Embryo Eye Sensory System
69477 69477 SRR18712326 SRX14812925 SRS12569568 SRP369458 PRJNA825495 Regulation of retina microexons by srrm3 in zebrafish PRJNA825495 Other To investigate the role of srrm3 as a regulator of retina microexons RetMICs we have sequenced heads from WT animals at 24 hpf and enucleated eyes from 48 hpf 60 hpf and 72 hpf larvae WT and KO for srrm3.         Eye 48hpf WT b   strain:WT|dev stage:48 hpf|sex:NA|tissue:Eye|BioSampleModel:Model organism or animal                 Eyes from 48 hpf larvae WT for the eMIC domain of srrm3. Eye 48hpf WT Eye 48hpf WT WT and srrm3 mutant zebrafish were grown at 28C 14h light/10h dark cycle. Extract protocol: 24 hpf WT fish heads were removed and pulled for RNA extraction using RNA RNeasy Mini kit QIAGEN. Larvae eyes were removed and from twenty to sixty eyes per genotype were pulled for RNA extraction using RNA RNeasy Micro kit QIAGENLibrary preparation: RNA libraries were prepared for sequencing using standard Illumina protocols; polyA selected stranded RNA seq     RNA-Seq TRANSCRIPTOMIC RANDOM PAIRED ILLUMINA Illumina HiSeq 2500   SRP369458     Eye_48hpf_WT_b_R1-125.fq.gz Eye_48hpf_WT_b_R2-125.fq.gz fastq fastq 15736041750.0 62944167.0 Eye 48hpf WT b R1 125.fq.gz 0:125 1:125 A:4076543478;C:3787178416;G:3834447099;T:4028745594;N:9127163 125 125     4076543478 3787178416 3834447099 4028745594 9127163 SRX14812925 SRS12569568 SRA1402039 Centre for Genomic Regulation|Systems Biology Centre for Genomic Regulation|Systems Biology 2 0.97095 0.97209 0.05101 0.05067 0.69408 0.69767 0.47501 0.47714 125 125 B B biological fallback assumption illumina hiseq_era unknown random_priming trueseq bulk unknown unknown   Spain 2022-04-11 Hatching Embryo Eye Sensory System
69478 69478 SRR18712327 SRX14812924 SRS12569567 SRP369458 PRJNA825495 Regulation of retina microexons by srrm3 in zebrafish PRJNA825495 Other To investigate the role of srrm3 as a regulator of retina microexons RetMICs we have sequenced heads from WT animals at 24 hpf and enucleated eyes from 48 hpf 60 hpf and 72 hpf larvae WT and KO for srrm3.         Eye 48hpf MUT b   strain:srrm3 MUT|dev stage:48 hpf|sex:NA|tissue:Eye|BioSampleModel:Model organism or animal                 Eyes from 48 hpf larvae mutated for the eMIC domain of srrm3. Eye 48hpf MUT Eye 48hpf MUT WT and srrm3 mutant zebrafish were grown at 28C 14h light/10h dark cycle. Extract protocol: 24 hpf WT fish heads were removed and pulled for RNA extraction using RNA RNeasy Mini kit QIAGEN. Larvae eyes were removed and from twenty to sixty eyes per genotype were pulled for RNA extraction using RNA RNeasy Micro kit QIAGENLibrary preparation: RNA libraries were prepared for sequencing using standard Illumina protocols; polyA selected stranded RNA seq     RNA-Seq TRANSCRIPTOMIC RANDOM PAIRED ILLUMINA Illumina HiSeq 2500   SRP369458     Eye_48hpf_MUT_b_R1-125.fq.gz Eye_48hpf_MUT_b_R2-125.fq.gz fastq fastq 14845531750.0 59382127.0 Eye 48hpf MUT b R1 125.fq.gz 0:125 1:125 A:3879709745;C:3539390575;G:3573173326;T:3844623832;N:8634272 125 125     3879709745 3539390575 3573173326 3844623832 8634272 SRX14812924 SRS12569567 SRA1402039 Centre for Genomic Regulation|Systems Biology Centre for Genomic Regulation|Systems Biology 2 0.9684 0.97006 0.05578 0.05498 0.69183 0.69581 0.472 0.46674 125 125 B B biological fallback assumption illumina hiseq_era unknown random_priming trueseq bulk unknown unknown   Spain 2022-04-11 Hatching Embryo Eye Sensory System

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