run_metadata: 32240

This data as json

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
32240	SRR29141378	SRX24663081	SRS21398379	SRP509393	PRJNA1115053	Large scale screen of novel zebrafish retinal ganglion cell ablation model reveals genetic regulation of retinal regeneration is context specific	GSE268179	Other	Many genes are known to regulate Müller glia MG dependent retinal regeneration following widespread tissue damage. Conversely genes controlling regeneration following limited retinal cell loss per degenerative disease are undefined. Studying regeneration in the context of selective cell loss is important as evidence suggests injury specifics inform the regenerative process. Here transgenic zebrafish enabling inducible selective retinal ganglion cell RGC ablation were combined with single cell multiomics and CRISPR/Cas9 based knockout methods to screen 101 genes for effects on RGC regeneration. We identified 18 regulators of RGC regeneration seven knockouts inhibited and eleven promoted RGC regeneration. Surprisingly 35 of 36 known/implicated regulators of retinal tissue regeneration following widespread damage were not required for RGC regeneration and seven of these knockouts actually enhanced RGC replacement kinetics including sox2 olig2 and ascl1a. Mechanistic analyses revealed ascl1a knockout increased the propensity of progenitor cells to produce RGCs; i.e. biased progenitor cell fate. These data demonstrate plasticity in how MG can convert to a stem like state and context specificity in how genes function during regeneration. Increased understanding of how disease relevant cell types can be selectively regenerated will support the development of disease tailored regenerative therapeutics. Overall design: We performed single cell RNA sequencing in larval zebrafish eyes following multiple paradigms of retinal damage including ablation of retinal ganglion cells RGCs 4 timepoints and ablation of rod photoreceptors and multiome sequencing following ablation of RGCs in fish with the ascl1a gene knocked out via CRISPR/Cas9.		pubmed:39007397		Unablated rgc:ntr day 9	GSM8287439		source name:Eye\|tissue:Eye\|transgenic line:rgc:ntr\|treatment:no mtz\|geo loc name:missing\|collection date:missing	Unablated rgc:ntr day 9	scRNAseq: Raw reads were mapped to the Danio rerio GRCz10 using Cell Ranger v7.0 from 10x genomics. Aligned genomic reads were then read into the published Seurat pipeline v4.3.0.1 and quality control was performed by removing any cells with <200 detected genes or 1000 UMIs and genes detected in fewer than 3 cells per experiment. Clustering steps were performed using steps from the pbmc Seurat tutorial available online. Briefly the top 2 000 variable genes were identified and used to identify principal components PCs of the data. The top 30 PCs were used to produce a UMAP and clusters were annotated with known zebrafish marker genes. Differentially expressed genes DEGs were identified using the FindAllMarkers function between each control and ablation timepoint in each retinal cell cluster minimum log2 foldchange cutoff of 0.25. scMultiomeseq: RNA expression data was processed as above. Peak calling from single nuclei ATAC seq reads was performed using MACS2 in the ArchR package v1.0.2. ATAC seq data was then processed using the pbmc scATAC seq workflow with the Signac v1.10.0 and Seurat v4.3.0.1 packages for quality control normalization and producing an integrated UMAP. Differential expression and accessibility was then calculated for both gene RNA expression and chromatin peak accessibility. Next the ChromVar package v1.18.0 was used to identify differentially accessible transcription factor motifs between wildtype and ascl1a mutant cells. Assembly: GRCz11 Supplementary files format and content: Cellular expression data varies in format either as h5 standalone files or barcodes features and matrix files to be used together. ATAC data is available as standalone fragment.tsv files	Eye		scRNAseq: 40 60 eyes were dissected from sibling fish and subsequently placed in 20 U/ml papain 10 eyes per 1 ml Worthington and incubated at 28°C for 30 min with gentle agitation. Cells were pelleted and resuspended in PBS containing 0.1 mg/ml leupeptin Sigma Aldrich and 10 U/ml DNaseI Roche. Cells were filtered through a 70 μm filter Miltenyi Biotec kept on ice until 10X genomics processing. scMultiomeseq: 40 60 eyes were dissected and flash frozen in dry ice for 15min before being transferred to a 80 C freezer for storage. Nuclei were extracted from frozen retinal tissues according to 10xMultiome ATAC + Gene Expression GEX protocol CGOOO338. Briefly frozen retinal tissues were lysed in ice cold 500ml of 0.1X Lysis buffer using a pestle and incubated on ice for 6 min totally. Nuclei were centrifuged washed 3 times and resuspended in 10xMultiome nuclei buffer at a concentration of 3000 5000 nuclei/ml and kept on ice until 10x genomics processing. Library preparation was then performed according to 10x genomics protocols.		tissue:Eye\|transgenic line:rgc:ntr\|treatment:no mtz	GSM8287439	GSM8287439: Unablated rgc:ntr day 9; Danio rerio; RNA Seq	GSM8287439 r1	GSM8287439	1	scRNAseq: 40 60 eyes were dissected from sibling fish and subsequently placed in 20 U/ml papain 10 eyes per 1 ml Worthington and incubated at 28°C for 30 min with gentle agitation. Cells were pelleted and resuspended in PBS containing 0.1 mg/ml leupeptin Sigma Aldrich and 10 U/ml DNaseI Roche. Cells were filtered through a 70 μm filter Miltenyi Biotec kept on ice until 10X genomics processing. scMultiomeseq: 40 60 eyes were dissected and flash frozen in dry ice for 15min before being transferred to a 80 C freezer for storage. Nuclei were extracted from frozen retinal tissues according to 10xMultiome ATAC + Gene Expression GEX protocol CGOOO338. Briefly frozen retinal tissues were lysed in ice cold 500ml of 0.1X Lysis buffer using a pestle and incubated on ice for 6 min totally. Nuclei were centrifuged washed 3 times and resuspended in 10xMultiome nuclei buffer at a concentration of 3000 5000 nuclei/ml and kept on ice until 10x genomics processing. Library preparation was then performed according to 10x genomics protocols.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000		SRP509393		loader:fastq load.py	TH134_S56_L003_R1_001.fastq.gz TH134_S56_L003_R2_001.fastq.gz	fastq fastq	4894210580.0	41127820.0	GSM8287439 r15	0:28 1:91	A:1389263901;C:1064650503;G:1124176385;T:1316003245;N:116546	28	91			1389263901	1064650503	1124176385	1316003245	116546	SRX24663081	SRS21398379	SRA1875751	Jeff Mumm, Ophthalmology, Johns Hopkins University	Jeff Mumm, Ophthalmology, Johns Hopkins University												T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x		United States	2024-05-23	Undetermined	Larval	Eye	Sensory System