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| Status |
Public on Apr 15, 2024 |
| Title |
wild-type P15 testes, biological replicate 2, scRNAseq |
| Sample type |
SRA |
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| Source name |
testes
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6N
tissue: testes
age: Postnatal day 15 (P15)
genotype: wild-type
batch: batch 2
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| Extracted molecule |
total RNA |
| Extraction protocol |
Single-cell sequencing libraries were prepared and sequenced in two batches, with each batch containing one P15 littermate pair comprised of one wild-type and one Meioc-null pup. One mouse testis per P15 pup was enzymatically dissociated by first shaking vigorously for 30 seconds and incubating at 35˚C for 7 minutes in 900 ul of 0.1% collagenase type I (Worthington LS004196) in Hank’s Balanced Salt Solution (HBSS), supplemented with TURBO DNase (ThermoFisher Scientific AM2238) for a final concentration of 2U/mL. After allowing the sample to sit for 1 minute such that the seminiferous tubules settled to the bottom of the tube, the interstitium was removed by discarding the resulting supernatant, and the remaining sample was incubated at 35˚C for 25 minutes (with gently pipetting every 5 minutes) in 800 ul of 0.05% trypsin and 0.1% collagenase type I in HBSS, supplemented with TURBO DNase for a final concentration of 2U/mL. The cell suspension was then passed through a 40um nylon strainer, and cell suspension was allowed to digest for 35˚C for an additional 20 minutes. The digestion was quenched by adding 200ul fetal bovine serum (FBS) and gently mixing. The cell suspension was again passed through a 40um nylon strainer. The concentration of the resulting cell suspension was assessed using trypan blue on the Countess Cell Counter. Cells were pelleted at 500g for 4 minutes at room temperature, resuspended in 0.05% bovine serum albumin (BSA) in phosphate buffered saline (PBS) for a target concentration of 1000 cells per microliter, and placed on ice. Viability and concentration were re-assessed ~20 minutes prior to loading on the Chromium Controller using trypan blue on a Countess Cell Counter, at which point all samples had a minimum of 90% viability. Cell suspensions were loaded onto the Chromium Controller, aiming for recovery of 10,000 cells per sample.
Libraries were generated using the Chromium Next GEM Single Cell 3ʹ v3.1 (10x Genomics), according to manufacturer’s instructions
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic single cell |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
10x Genomics
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| Data processing |
Alignment, filtering, barcode counting, and UMI counting were done via count function in Cell Ranger v.4.0.0 with default settings using Cell Ranger’s mm10-2020-A reference package (i.e., the GRCm38/mm10 mouse genome assembly with GENCODE vM23/Ensembl 98 annotation)
Assembly: mm10
Supplementary files format and content: Tab-separated values files and matrix files.
Supplementary files format and content: rds file contains the processed data for all 4 samples from GSE230746 as a seurat object containing the count matrix, metadata, clustering results, and labeled cluster cell types.
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| Submission date |
Apr 27, 2023 |
| Last update date |
Apr 15, 2024 |
| Contact name |
Maria M. Mikedis |
| E-mail(s) |
maria.mikedis@cchmc.org
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| Organization name |
Cincinnati Children's Hospital Medical Center
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| Street address |
240 Albert Sabin Way
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| City |
Cincinnati |
| State/province |
OH |
| ZIP/Postal code |
45229 |
| Country |
USA |
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| Platform ID |
GPL24247 |
| Series (2) |
| GSE230746 |
The role of MEIOC at the transition from mitosis to meiosis during mammalian spermatogenesis [scRNA] |
| GSE230747 |
The role of MEIOC at the transition from mitosis to meiosis during mammalian spermatogenesis |
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| Relations |
| BioSample |
SAMN34403537 |
| SRA |
SRX20117524 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM7233605_WT_P15_testis_rep2_barcodes.tsv.gz |
70.4 Kb |
(ftp)(http) |
TSV |
| GSM7233605_WT_P15_testis_rep2_features.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSM7233605_WT_P15_testis_rep2_matrix.mtx.gz |
161.2 Mb |
(ftp)(http) |
MTX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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