run_metadata: 56714

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
56714	SRR11040632	SRX7692415	SRS6118509	SRP247644	PRJNA605419	FGFR3 is a positive regulator of osteoblast expansion and differentiation during zebrafish skull vault development	GSE144944	Transcriptome Analysis	In order to study FGFRthree primes role in cranial vault development we generated the first fgfr3 loss of function zebrafish fgfr3lof1/lof1. The mutant fish exhibited major changes in the craniofacial skeleton with a lack of sutures abnormal frontal and parietal bones and the presence of ectopic bones. In order to define cellular and molecular processes responsible for this phenotype we performed single cell RNA sequencing of cranial vault cells from fgfr3lof1/lof1 and fgfr3+/+ zebrafish. Our data allow us to define transcriptomic profile of each osteogenic subpopulation involved in cranial vault formation and we revealed a defect in osteoblast differentiation associated with changes in the extracellular matrix. These findings demonstrate that fgfr3 is a positive regulator of osteogenesis and we conclude that changes in the extracellular matrix within growing bone impair cell cell communication mineralization and new osteoblast recruitment. Overall design: Using cells isolated from cranial vaults from two fgfr3lof1/lof1 fish and two fgfr3+/+ zebrafish fgfr3lof/lof: zebrafish line was obtained using CrisPR/Cas9 technology with a stop codon at position 377 we profiled the transcriptome of nearly 28000 single cells using the Chromium system 10x Genomics. Cells suspensions were obtained using both collagenase digestion and cell sorting to eliminate death cells. Cells of the cranial vault is a mix of several cell types osteogenic cells chondrogenic cells immune cells epidermal cells endothelial cells. The scRNA seq libraries were generated with a Chromium Single Cell three prime Library & Gel Bead Kit v.3 10x Genomics. The purified libraries were sequenced on a Novaseq 6000 Illumina on paired end strands read 1: 28bp; read 2: 91bp with a mean read depth of 25000 reads per cell.		pubmed:32379366		2 fgfr3+/+	GSM4301249		source name:Cranial vault\|tissue:Cranial vault\|genotype/variation:fgfr3+/+\|length:9 mm	2 fgfr3+/+	Fastq files from the scRNA 10X libraries were processed using the Cell Ranger Count pipeline with its default parameters. Reads were aligned against the GRCz11 3.0.0 reference transcriptome. The RNA data quality control and downstream analysis were performed using the Seurat R package version 3.0.2 and the standard Seurat v3 integration workflow. We filtered cells that had i unique feature counts over 2000 or below 500 or ii more than 20% of mitochondrial counts. post data filtering we obtained an expression matrix with 21176 genes and 19245 cells. The matrix’s dimensions were reduced by running the 20 significant principal components against scaled data. Corresponding to an object of class Seurat SC fgfr3 Cranial Vault ZF combined samples. We used the FindMarkers function to define marker of each cluster Findmarkers by cluster.xlsx file. We used the FindMarkers function to compute differentially expressed genes DEGs for each of the identity classes in the dataset DEGS.xlsx file. We subset cluster 4 from SC fgfr3 Cranial Vault ZF combined samples matrix and performed new clustering resolution 0.2 and we obtained SC fgfr3 Cranial Vault ZF Subset Cluster4. Genome build: GRCz11 3.0.0 Supplementary files format and content: * raw feature bc matrix.h5 Supplementary files format and content: Findmarkers by cluster.xlsx Supplementary files format and content: DEGS.xlsx	Cranial vault		The cranial vault was isolated from the head of 9SL zebrafish and dissected in PBS. The olfactory bulbs the anterior parts of the frontal bones and the suboccipital bones were removed. The soft tissue was partially degraded by incubation with 0.2% collagenase and 4 mM EDTA in PBS for 10 min at 28°C. post a wash in PBS cells were dissociated for 40 min at 28°C in 0.2% collagenase in PBS. The solution was strained using a 40 μm cell strainer and the cells were washed in a MEM supplement with 10% FBS. Prior to flow cytometry the cells were suspended in FacsMax and stained with SytoxTM green nucleic acid stain and Hoechst. To sort the cells beads were used to select events larger than 3 µm. Hoechst positive cells cells with a nucleus and Syber green negative cells live cells were sorted. The scRNA seq libraries were generated with a Chromium Single Cell 3′ Library & Gel Bead Kit v.3 10x Genomics according to the manufacturer’s instructions. Briefly cells were counted diluted at 1000 cells/µL in PBS+0.04% BSA and 6500 cells were loaded in the 10x Chromium Controller to generate single cell gel beads in an emulsion. post reverse transcription the gel beads in the emulsion were disrupted. Barcoded complementary DNA was isolated and amplified in PCRs. Following fragmentation end repair and A tailing sample indexes were added during index PCR. The purified libraries were sequenced on a Novaseq 6000 Illumina on paired end strands read 1: 28bp; read 2: 91bp with a mean read depth of 25000 reads per cell.		tissue:Cranial vault\|genotype/variation:fgfr3+/+\|length:9 mm	GSM4301249	GSM4301249: 2 fgfr3+/+; Danio rerio; RNA Seq	GSM4301249		1	The cranial vault was isolated from the head of 9SL zebrafish and dissected in PBS. The olfactory bulbs the anterior parts of the frontal bones and the suboccipital bones were removed. The soft tissue was partially degraded by incubation with 0.2% collagenase and 4 mM EDTA in PBS for 10 min at 28°C. post a wash in PBS cells were dissociated for 40 min at 28°C in 0.2% collagenase in PBS. The solution was strained using a 40 μm cell strainer and the cells were washed in a MEM supplement with 10% FBS. Prior to flow cytometry the cells were suspended in FacsMax and stained with SytoxTM green nucleic acid stain and Hoechst. To sort the cells beads were used to select events larger than 3 µm. Hoechst positive cells cells with a nucleus and Syber green negative cells live cells were sorted. The scRNA seq libraries were generated with a Chromium Single Cell 3′ Library & Gel Bead Kit v.3 10x Genomics according to the manufacturer's instructions. Briefly cells were counted diluted at 1000 cells/µL in PBS+0.04% BSA and 6500 cells were loaded in the 10x Chromium Controller to generate single cell gel beads in an emulsion. post reverse transcription the gel beads in the emulsion were disrupted. Barcoded complementary DNA was isolated and amplified in PCRs. Following fragmentation end repair and A tailing sample indexes were added during index PCR. The purified libraries were sequenced on a Novaseq 6000 Illumina on paired end strands read 1: 28bp; read 2: 91bp with a mean read depth of 25000 reads per cell.	GEO Accession:GSM4301249	RNA-Seq	TRANSCRIPTOMIC	cDNA	PAIRED	ILLUMINA	Illumina NovaSeq 6000		SRP247644			2-fgfr3plus-R3-ZF-0519_S6_L001_R1_001.fastq.gz 2-fgfr3plus-R3-ZF-0519_S6_L001_R2_001.fastq.gz	fastq fastq	4962416501.0	41700979.0	GSM4301249 r1	0:28 1:91	A:1356709063;C:1139750321;G:1170875983;T:1294727572;N:353562	28	91			1356709063	1139750321	1170875983	1294727572	353562	SRX7692415	SRS6118509	SRA1039424	GEO	Molecular and physiopathological bases of osteochondrodysplasia, U1163, Institut Imagine	2	0.00727	0.95235	0.00248	0.13046	0.98981	0.78054	0.33659	0.48621	28	91	T	B	sc-like readlen	illumina	novaseq_era	unknown	cdna_unspecified	trueseq	sc	single_cell_droplet	10x		France	2020-02-07	Undetermined	Undetermined	Multi-tissue	Multi-system