run_metadata: 65065

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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
65065	SRR14923995	SRX11237267	SRS9283141	SRP325759	PRJNA741770	Single cell profiling of cranial neural crest diversification across a vertebrate lifetime	GSE178969	Other	The cranial neural crest CNC is a vertebrate specific population that generates a huge diversity of derivatives including the bulk of the connective and skeletal tissues of the head. How neural crest cells generate the appropriate cell types for distinct head regions over time remains unresolved. Here we profile RNA expression scRNAseq and chromatin accessibility snATACseq of the zebrafish cranial neural crest lineage at single cell resolution from embryos to adults. Overall design: To recover the cell heterogeneity and lineage information in the span of a life time of a fish we collected the converted cells from embryonic stages 1.5 2 dpf larval stage 3 5 and 14 dpf juvenile stage 60 dpf to adult stage 150 or 210 dpf. We used a cranial neural crest converted zebrafish line 28.5Sox10:Cre;bactin:BFP to DsRed to permanantly convert and label the CNCC derived cells. The converted cells were collected and subjected to single cell sequencing for both their transcriptome and open chromatin region using Chromium platform of 10X Genomic. For biological and technical control we include replicates for both scRNAseq and snATACseq at various stages. We include an additional line fli1a:eGFP/sox10:dsRed for a biological and positive control of CNCC of 1.5 dpf sample.		pubmed:35013168		scRNAseq Sox10 Cre bact BtR 60dpf rep 1	GSM5402439		tissue:Cranial neural crest derived cells\|cell type:Cranial neural crest derived cells\|transgene1:TgMmu.Sox10 Mmu.Fos:Crezf384\|transgene2:Tgactab2:loxP BFP STOP loxP dsRed\|developmental stage:60 dpf\|facs markers:Zombie / dsRed+	scRNAseq Sox10 Cre bact BtR 60dpf rep 1	For scRNAseq libraries Cellranger v3.0.0 10X Genomics was used for demultiplexing alignment and raw count matrix generation. The alignment was performed against zebrafish genome index built with GRCz11.fa and GRCz11.98.gtf. For snATACseq libraries Cellranger ATAC v1.2.0 10X Genomics was used for alignment against zebrafish genome index built with GRCz11.fa JASPAR2020 and GRCz11.98.gtf peak calling and peak by cell count matrix generation with default parameters. QC step for the library was performed along with the Cellranger or Cellranger ATAC alignment process. We performed a second round of peak calling for our snATACseq data using package Snaptools and SnapATAC. We first removed cell debris low quality cells and cell duplicates by setting cutoffs based on percentage of reads in peaks > 30 for 60 dpf > 45 for 210 dpf and > 50 for the rest and fragment number within peaks 5 000 – 30 000 for 5 dpf 1 000 – 11 000 for 14 dpf and 1 000 – 20 000 for the rest. To recover every aligned fragment we binned the genome into 5 kb sections and constructed the bin by cell matrices bmats for each library by Snaptools from the positional sorted bam files generated by cellranger atac. The cells were filtered dimensionally reduced by diffusion map and clustered with inputs of the first 34 dimensions followed the SnapATAC vignette. The specific peaks were called for each cluster by the wrapped MACS2 function in SnapATAC with parameter gsize = 1.5e9 shift = 100 ext = 200 and qval = 5e 2. The finalized and refined peak profile was derived by collapsing and merging all 175 individual peak files to about 450K peaks. The count matrices of both scRNAseq and snATACseq data were analyzed by R package Seurat and Signac. The count matrices of each sample were aggregated if there is any replicate. For scRNAseq data the matrices are normalized NormalizeData and scaled for the top 2 000 variable genes FindVariableFeatures and ScaleData. The scaled matrices were dimensional reduced to 50 principal components 60 components for 150 dpf which then subjected to neighbor finding FindNeighbors k = 20 and clustering FindClusters resolution = 0.8. The data were visualized through UMAP with 50 principal components as input. For snATACseq data the matrices are dimensionally reduced 30 latent semantic indices LSIs through RunTFIDF and RunSVD functions. The neighbor finding clustering and visualization are performed with the second to the thirtieth LSIs as input algoritm = 3 for FindClusters. Genome build: GRCz11 Supplementary files format and content: Each sample comes with four processed data files: a tab delimited cell barcode file tsv a tab delimited feature file with genes for scRNAseq or peaks for snATACseq tsv a sparse matrix of count table mtx and a cluster table tsv. The previous three files are suffice for creating a single cell count matrix. The cluster table can be further added to recreat the corresponding cluster information as published.	Cranial neural crest derived cells		Dissected heads from converted Sox10:cre; bactin:loxP eGFP loxP DsRed fish were incubated in fresh Ringer’s solution 5 10 min followed by mechanical and enzymatic dissociation by pipetting every 5 minutes in protease solution 0.25% trypsin Life Technologies 15090 046 1 mM EDTA and 400 mg/ml Collagenase D Sigma 11088882001 in PBS and incubated at 28.5°C for 20 30 minutes or until full dissociation. Reaction was stopped by 6X stop solution 6 mM CaCl2 and 30% fetal bovine serum FBS in PBS. Cells were pelleted 2000 rpm 5 minute 4 °C and resuspended in suspension media 1% FBS 0.8 mM CaCl2 50 U/ml penicillin and 0.05 mg/ml streptomycin Sigma Aldrich St. Louis MO in phenol red free Leibovitz’s L15 medium Life Technologies twice. Final volumes of 500 µl resuspended cells were placed on ice and sorted by fluorescence activated cell sorting FACS to isolate live cells that excluded the cytoplasmic stain Zombie green BioLegend 423111 or the nuclear stain DAPI. Live cells were sorted into PBS solution containing 0.04% BSA and loaded to 10X Genomics microfluidic chips. The version 2 of Chromium Single Cell three prime Reagent Kits was used for RNA extraction. For snATACseq library construction we used the same cell dissociation and sorting protocol as for scRNAseq with isolation of live cells that excluded the cytoplasmic stain Zombie green product information. Nuclei isolation was performed per manufacturer’s instructions 10X Genomic protocol CG000169. Cells were incubated with lysis buffer on ice for 90 second followed by integrity check of nuclei under fluorescence microscope with DAPI before library synthesis. For scRNAseq the cell lysis polyA pull down tagging with cell barcode and RNA:DNA hybrid synthesis were performed in the GEMs per manufacturer's instruction. GEMs containing barcoded single cell cDNA were lysis for further fragmentation and ligation of illumina P5 and P7 ends. Libraries were sequenced to depths of at least 1 000 000 reads per cell for each library. For snATACseq the open chromatins of the isolated nuclei were targeted and tagged by transposase barcoded and ligated with Illumina P5 sequence in the individually encapsulated GEMs per manufacturer's instruction. Barcoded single nucleus genomic DNA were released from GEMs and pulled down for further ligation of library specific index and Illumina P7 sequence. Libraries were sequenced to depths of at least 75 000 read pairs per nucleus for each library.	The Institutional Animal Care and Use Committee of the University of Southern California approved all animal experiments Protocol 20771. Zebrafish embryos Danio rerio were raised in embryo media at 28.5°C. For adults in the institute vivarium according to Protocol 20771.	cell type:Cranial neural crest derived cells\|transgene1:TgMmu.Sox10 Mmu.Fos:Crezf384\|transgene2:Tgactab2:loxP BFP STOP loxP dsRed\|developmental stage:60 dpf\|facs markers:Zombie / dsRed+	GSM5402439	GSM5402439: scRNAseq Sox10 Cre bact BtR 60dpf rep 1; Danio rerio; RNA Seq	GSM5402439		1	Dissected heads from converted Sox10:cre; bactin:loxP eGFP loxP DsRed fish were incubated in fresh Ringer's solution 5 10 min followed by mechanical and enzymatic dissociation by pipetting every 5 minutes in protease solution 0.25% trypsin Life Technologies 15090 046 1 mM EDTA and 400 mg/ml Collagenase D Sigma 11088882001 in PBS and incubated at 28.5°C for 20 30 minutes or until full dissociation. Reaction was stopped by 6X stop solution 6 mM CaCl2 and 30% fetal bovine serum FBS in PBS. Cells were pelleted 2000 rpm 5 minute 4 °C and resuspended in suspension media 1% FBS 0.8 mM CaCl2 50 U/ml penicillin and 0.05 mg/ml streptomycin Sigma Aldrich St. Louis MO in phenol red free Leibovitz's L15 medium Life Technologies twice. Final volumes of 500 µl resuspended cells were placed on ice and sorted by fluorescence activated cell sorting FACS to isolate live cells that excluded the cytoplasmic stain Zombie green BioLegend 423111 or the nuclear stain DAPI. Live cells were sorted into PBS solution containing 0.04% BSA and loaded to 10X Genomics microfluidic chips. The version 2 of Chromium Single Cell three prime Reagent Kits was used for RNA extraction. For snATACseq library construction we used the same cell dissociation and sorting protocol as for scRNAseq with isolation of live cells that excluded the cytoplasmic stain Zombie green product information. Nuclei isolation was performed per manufacturer's instructions 10X Genomic protocol CG000169. Cells were incubated with lysis buffer on ice for 90 second followed by integrity check of nuclei under fluorescence microscope with DAPI before library synthesis. For scRNAseq the cell lysis polyA pull down tagging with cell barcode and RNA:DNA hybrid synthesis were performed in the GEMs per manufacturer's instruction. GEMs containing barcoded single cell cDNA were lysis for further fragmentation and ligation of illumina P5 and P7 ends. Libraries were sequenced to depths of at least 1 000 000 reads per cell for each library. For snATACseq the open chromatins of the isolated nuclei were targeted and tagged by transposase barcoded and ligated with Illumina P5 sequence in the individually encapsulated GEMs per manufacturer's instruction. Barcoded single nucleus genomic DNA were released from GEMs and pulled down for further ligation of library specific index and Illumina P7 sequence. Libraries were sequenced to depths of at least 75 000 read pairs per nucleus for each library.	GEO Accession:GSM5402439	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina HiSeq 4000		SRP325759			scRNAseq_Sox10_Cre_bact_BtR_60dpf_rep_1_S14_L001_R1_001.fastq.gz scRNAseq_Sox10_Cre_bact_BtR_60dpf_rep_1_S14_L001_R2_001.fastq.gz	fastq fastq	7897275260.0	53567183.0	GSM5402439 r2	0:27 1:120.43	A:2323096260;C:1712849172;G:1740350109;T:2116414828;N:4564891	27	120			2323096260	1712849172	1740350109	2116414828	4564891	SRX11237267	SRS9283141	SRA1250944	GEO	Crump Lab, Stem Cell, USC	2	0.00136	0.85419	0.00056	0.13524	0.99709	0.85989	0.38853	0.51984	27	120	T	B	sc-like readlen	illumina	hiseq_era	unknown	poly_a	unknown	sc	single_cell_droplet	10x		United States	2021-06-27	Juvenile	Juvenile	Brain	Nervous System