run_metadata: 75487

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
75487	SRR24641286	SRX20421318	SRS17735853	SRP438460	PRJNA973994	sc RNAseq of the day 4 zebrafish spinal cord	GSE232801	Transcriptome Analysis	Identification of the neuronal types that form the specialized circuits controlling distinct behaviors has benefited greatly from the simplicity offered by zebrafish. Electrophysiological studies have shown that additional to connectivity understanding of circuitry requires identification of functional specializations among individual circuit components such as those that regulate levels of transmitter release and neuronal excitability. In this study we use single cell RNAseq scRNAseq to identify molecular distinctions causal to the unique physiology of primary motoneuron PMn function as well as associated specialized interneurons that are tailored specifically for mediation of the powerful escape response. Generating scRNA seq datasets that provide both a wide overview of the cellular populations of the spinal cord and a granular view of the zebrafish motor neurons. Overall design: Cells from the spinal cords of 150 tgSAIG213a;EGFP zebrafish were analyzed via scRNA seq and populations of interneurons were identified based on established markers. For examination of Motor neurons we additionally preformed FACS on cells dissociated from spinal cords of 150 tgmnx1:GFP zebrafish.		pubmed:37333232;pubmed:37975797		Spinal cord saig rep 1	GSM7383490		source name:spine\|tissue:spine\|cell type:mixed spinal cells\|genotype:TgSAIG213A;EGFP\|developmental stage:4 dpf\|geo loc name:missing\|collection date:missing	Spinal cord saig rep 1	Barcode processing and gene counting were preformed using Cell ranger software v6.1.1 10X Genomics. Reads were aligned to a reference genome made using cellrangers mkref command using the GRCz11.fa and the zebrafish custom ref.GTF provided as a supplimental file. The zebrafish custom ref.GTF is a lightly modified version of the Lawson v 1.4.3 annotation Lawson et al 2020 eLife in which GFP has been added and a number of naming mistakes have been corrected. Normalization integration and quality control were preformed using Seurat. Clustering of the data was preformed using the Lieden algorithm implemented in Seurat with varying parameters depending on the sample. Clusters with markers indicating contamination hindbrain and muscle were removed as were clusters where over 90% of cells came from only one experimental run. Neurons and glia were isolated based on the expression of markers within clusters snap25a and elavl4 for neurons and slc1a2b gfap myrf and sox10 for glia. Neurons were then sub clustered into 33 clusters and identities were assigned to 21 of them based on both established markers in the literature and a number of novel markers we were able to validate. Motor neurons were isolated in the dataset based on the expression of motor neuron markers mnx1 mnx2b isl1 and cholinergic markers slc18a3a and chata within clusters. Motor neurons isolated this way were then integrated together using Seurat. Assembly: GRCz11 Supplementary files format and content: Tab seperated value files and matrix files providing the processed count matrices for each sample Supplementary files format and content: RDS file providing the Seurat data object for the integrated spinal samples Supplementary files format and content: RDS file providing the Seurat data object for the annotated set of clustered neurons from the integrated spinal samples Supplementary files format and content: RDS file providing the Seurat data object for the integrated set of isolated motor neurons from all samples Supplementary files format and content: GTF format record of the reference genome used in the creation of this data set	spine		About 150 dpf 4 dpf larva were euthanized in 0.02% tricaine and individually decapitated behind the hindbrain. They were incubated with 20 mg/ml collagenase in bath solution 134 mM NaCl 2.9 mM KCl 1.2 mM MgCl2 2.1 mM CaCl2 and 10 mM Na HEPES pH 7.8 ∼290 mOsm at 28 °C for 1.5 hr with intermittent trituration using a p200 pipette aid at 0 0.5 hr and1 hr of the incubation. For the purpose of isolating spinal cords from remaining tissue the final trituration was done using fire polished Pasteur pipettes with decreased opening sizes 300 200 100 μm respectively. Intact spinal cords were transferred to L15 media and washed 3 times with fresh media. The spinal cords were incubated with 0.25% trypsin solution in 1xPBS containing 1 mM EDTA at 28 °C for 25 min. The digestion was terminated by adding 500 μl stop solution L15 with 1% fetal bovine serum. The tissue was collected by spinning at 400 g for 3 min at 4°C washed once with L15 and resuspended in 200 μl of L15 media. Spinal cord cells were dissociated by triturating the digested tissue with fire polished Pasteur pipettes with 80 100 μm opening. The solution was filtered through a 35μm strainer into a siliconized collection tube. The suspension was examined on a microscope for cell count Trypan blue staining based viability test and proportion of dispersed single cells. Samples with a viability above 70% were used for sequencing. For FACS sorted samples single cell suspension was prepared as above from Tgmnx1:GFP fish and were FAC sorted for EGFP+ cells using a 100 μm nozzle on a BD inFlux cell sorter Flow Cytometry Shared Resource OHSU Single cell capture cDNA synthesis and library preparation were performed by the Massive Parallel Sequencing Shared Resource at OHSU using the 10x Genomics Chromium v3.0 reagent kit. Single cell suspension for the two full spinal cord replicates targeted 10 000 15 000 cells for the two FACS sorted samples 4 000 5 000 cells were targeted.		tissue:spine\|cell type:mixed spinal cells\|genotype:TgSAIG213A;EGFP\|developmental stage:4 dpf	GSM7383490	GSM7383490: Spinal cord saig rep 1; Danio rerio; RNA Seq	GSM7383490 r1	GSM7383490	1	About 150 dpf 4 dpf larva were euthanized in 0.02% tricaine and individually decapitated behind the hindbrain. They were incubated with 20 mg/ml collagenase in bath solution 134 mM NaCl 2.9 mM KCl 1.2 mM MgCl2 2.1 mM CaCl2 and 10 mM Na HEPES pH 7.8 ∼290 mOsm at 28 °C for 1.5 hr with intermittent trituration using a p200 pipette aid at 0 0.5 hr and1 hr of the incubation. For the purpose of isolating spinal cords from remaining tissue the final trituration was done using fire polished Pasteur pipettes with decreased opening sizes 300 200 100 μm respectively. Intact spinal cords were transferred to L15 media and washed 3 times with fresh media. The spinal cords were incubated with 0.25% trypsin solution in 1xPBS containing 1 mM EDTA at 28 °C for 25 min. The digestion was terminated by adding 500 μl stop solution L15 with 1% fetal bovine serum. The tissue was collected by spinning at 400 g for 3 min at 4°C washed once with L15 and resuspended in 200 μl of L15 media. Spinal cord cells were dissociated by triturating the digested tissue with fire polished Pasteur pipettes with 80 100 μm opening. The solution was filtered through a 35μm strainer into a siliconized collection tube. The suspension was examined on a microscope for cell count Trypan blue staining based viability test and proportion of dispersed single cells. Samples with a viability above 70% were used for sequencing. For FACS sorted samples single cell suspension was prepared as above from Tgmnx1:GFP fish and were FAC sorted for EGFP+ cells using a 100 μm nozzle on a BD inFlux cell sorter Flow Cytometry Shared Resource OHSU Single cell capture cDNA synthesis and library preparation were performed by the Massive Parallel Sequencing Shared Resource at OHSU using the 10x Genomics Chromium v3.0 reagent kit. Single cell suspension for the two full spinal cord replicates targeted 10 000 15 000 cells for the two FACS sorted samples 4 000 5 000 cells were targeted.		RNA-Seq	TRANSCRIPTOMIC SINGLE CELL	cDNA	PAIRED	ILLUMINA	NextSeq 500		SRP438460		loader:fastq load.py	CEL210928PB_HW1011_SAIG_D4_S2_L002_I1_001.fastq.gz CEL210928PB_HW1011_SAIG_D4_S2_L002_I2_001.fastq.gz CEL210928PB_HW1011_SAIG_D4_S2_L002_R1_001.fastq.gz CEL210928PB_HW1011_SAIG_D4_S2_L002_R2_001.fastq.gz	fastq fastq fastq fastq	76811238680.0	349141994.0	GSM7383490 r2	0:10 1:10 2:100 3:100	A:18612770086;C:13983377544;G:14583499379;T:22647109701;N:1642090	10	10	100	100	18612770086	13983377544	14583499379	22647109701	1642090	SRX20421318	SRS17735853	SRA1640659	Brehm, Vollum, Oregon Health and Science University	Brehm, Vollum, Oregon Health and Science University	2	9e-05	0.87854	0.0	0.2305	0.99997	0.81793	0.0	0.65388	100	100	T	B	mate1 technical by mapping diff	illumina	nextseq	unknown	cdna_unspecified	unknown	sc	single_cell_droplet	10x		United States	2023-05-18	Larval	Larval	Multi-tissue	Multi-system