{"database": "metadata", "table": "run_metadata", "rows": [[65100, "SRR14923958", "SRX11237258", "SRS9283133", "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.", null, "pubmed:35013168", null, "scRNAseq Sox10 Cre bact BtR 2dpf rep 1", "GSM5402430", null, "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:2 dpf|facs markers:Zombie  / dsRed+", "scRNAseq Sox10 Cre bact BtR 2dpf 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 \u2013 30 000 for 5 dpf  1 000 \u2013 11 000 for 14 dpf  and 1 000 \u2013 20 000 for the rest.  To\u00a0 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", null, "Dissected heads from converted Sox10:cre; bactin:loxP eGFP loxP DsRed fish were incubated in fresh Ringer\u2019s 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\u00b0C 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 \u00b0C 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\u2019s L15 medium Life Technologies twice. Final volumes of 500 \u00b5l 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\u2019s 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\u00b0C. 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:2 dpf|facs markers:Zombie  / dsRed+", "GSM5402430", "GSM5402430: scRNAseq Sox10 Cre bact BtR 2dpf rep 1; Danio rerio; RNA Seq", "GSM5402430", null, "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\u00b0C 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 \u00b0C 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 \u00b5l 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:GSM5402430", "RNA-Seq", "TRANSCRIPTOMIC", "cDNA", "PAIRED", "ILLUMINA", "Illumina HiSeq 4000", null, "SRP325759", null, null, "scRNAseq_Sox10_Cre_bact_BtR_2dpf_rep_1_S1_L001_R1_001.fastq.gz scRNAseq_Sox10_Cre_bact_BtR_2dpf_rep_1_S1_L001_R2_001.fastq.gz", "fastq fastq", 34509011947.0, 234007853.0, "GSM5402430 r1", "0:27 1:120.47", "A:9735364598;C:7434652978;G:7783819912;T:9534313520;N:20860939", 27, 120, null, null, 9735364598, 7434652978, 7783819912, 9534313520, 20860939, "SRX11237258", "SRS9283133", "SRA1250944", "GEO", "Crump Lab, Stem Cell, USC", 2, 0.0067, 0.90889, 0.00161, 0.09851, 0.98518, 0.8173, 0.48738, 0.49248, 27, 120, "T", "B", "sc-like readlen", "illumina", "hiseq_era", "unknown", "poly_a", "unknown", "sc", "single_cell_droplet", "10x", null, "United States", "2021-06-27", "Hatching", "Embryo", "Brain", "Nervous System"]], "columns": ["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"], "primary_keys": ["rowid"], "primary_key_values": ["65100"], "units": {}, "query_ms": 11.219122003240045}