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SRX4112602: GSM3149744: Cochlea SGNs Mouse 2; Mus musculus; RNA-Seq
2232 ILLUMINA (Illumina NovaSeq 6000) runs: 86.4M spots, 6.5G bases, 3.5Gb downloads

Submitted by: NCBI (GEO)
Study: Transcriptional profiling of cochlear neurons
show Abstracthide Abstract
In order to characterize the various subpopulations of neurons present in the inner ear, we dissected out the cochleas from wild-type mice, removing non-neuronal populations as possible, then dissociated the tissue to obtain a single cell suspension and profiled these by single cell RNA-Seq. Overall design: This study''s goal was to characterize the native populations of murine cochlear neurons. To ensure reproducibility of our findings, neurons from 4 separate mice were profiled.
Sample: Cochlea SGNs Mouse 2
SAMN09237862 • SRS3328056 • All experiments • All runs
Organism: Mus musculus
Library:
Instrument: Illumina NovaSeq 6000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: cDNA
Layout: SINGLE
Construction protocol: Temporal bones from 9 wko, male CBA/CaJ mice (n = 8 capsules from 4 mice) were isolated in ice-cold Leibovitz's L-15 medium, and the overlying bone and lateral wall was extracted from apex to base, leaving just the modiolus. The remaining structures consisted of the spiral ganglion, the spiral limbus, inner sulcus, and portions of the organ of Corti and outer sulcus. Microdissected tissue from each mouse was pooled in a solution of Leibovitz containing 200 units/mL collagenase IV (Sigma) and 10 kunitz/ml DNase I (Stem Cell Technologies) and incubated for 30 min at 37 °C to digest extracellular matrix. The collagenase IV solution was then replaced with EBSS containing 100 units of papain (Worthington Biochemical), 1.6 mM L-cysteine, 0.8 mM EDTA, 15 mM HEPES, and 10 kunitz/ml DNase I and incubated for an additional 30 min at 37 °C, triturating with a 1000 uL pipette every 10 min to generate a single cell suspension. An equal volume of Leibovitz containing 20% ovomucoid protease inhibitor (Worthington Biochemical) was added, and the dissociated cells were passed through a 20 mm filter (pluriSelect) to remove large debris. The cells were pelleted at 300g for 5 min, washed with PBS, and then pelleted and resuspended in PBS containing 0.1% BSA. Finally, a 10 uL sample of the cell suspension was counted on a Luna Fl automated counter using a Live/Dead assay (Thermo Fisher Scientific). Total time from euthanasia to single-cell capture was 2.5 h. The cell suspension was diluted to a concentration of ~10,000 cells per uL and immediately captured, lysed, and primed for reverse transcription (RT) using the high throughput, droplet microfluidics Gemcode platform from 10X Genomics with v2 chemistry (Zheng et al., 2017). Each droplet on the Gemcode co-encapsulates a cell and a gel bead that is hybridized with oligo(dT) primers encoding a unique cell barcode and unique molecular identifiers (UMIs) in lysis buffer. The capture process takes 5 min, after which the transcriptomes captured on gel beads are pooled and immediately reverse transcribed to cDNA. Since all cDNA is pooled and PCR amplified, cell barcodes and UMIs facilitate demultiplexing of the originating cell and mRNA transcript after sequencing. RT, PCR amplification of cDNA, and preparation of a library from 3' ends were conducted according to the manufacturer's published protocol. The library was sequenced on an Illumina NovaSeq 6000 at the Broad Institute using 2 x 100 bp paired-end reads.
Experiment attributes:
GEO Accession: GSM3149744
Links:
Runs: 2,232 runs, 86.4M spots, 6.5G bases, 3.5Gb
Run# of Spots# of BasesSizePublished
SRR719801337,7662.8M1.8Mb2018-08-14
SRR719801436,7162.8M1.3Mb2018-08-14
SRR719801540,3503M2.1Mb2018-08-14
SRR719801629,2382.2M1.4Mb2018-08-14
SRR719801727,7242.1M1.2Mb2018-08-14
SRR719801811,687876,525790,226b2018-08-14
SRR719801912,554941,550706,373b2018-08-14
SRR719802013,122984,150756,869b2018-08-14
SRR719802113,4031M884,679b2018-08-14
SRR719802221,8981.6M1.1Mb2018-08-14
SRR719802323,7381.8M1.1Mb2018-08-14
SRR71980247,930594,750753,477b2018-08-14
SRR719802515,5871.2M857,029b2018-08-14
SRR719802632,9412.5M1.6Mb2018-08-14
SRR719802722,4331.7M1,035,276b2018-08-14
There are 2,217 omitted runs. See all runs in Run Selector.

ID:
5598163

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