|
| Status |
Public on Oct 21, 2019 |
| Title |
KO-6 |
| Sample type |
SRA |
| |
|
| Source name |
testis
|
| Organism |
Mus musculus |
| Characteristics |
strain: B6D2F1 inbred
tissue: testis
age: post natal day 7
genotype: Pramef12-/-
|
| Growth protocol |
B6D2F1 mice were housed in NIDDK mouse core facility, in accordance with guidelines of the Animal Care and Use Committee of the National Institutes of Health under a Division of Intramural Research, NIDDK approved animal study protocols. Testes isolated from P2 or P7 aged control and knockout mice.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
Testes were isolated, total RNA was isolated using RNeasy Mini Kit and mRNA was purified by Dynabeads mRNA Purification Kit.
First strand cDNA was synthesized with SuperScript III Reverse Transcriptase. For second strand cDNA synthesis, the 100 μl reaction system included: 20 μl of first strand cDNA synthesis mix, 10 μl of 10x second strand buffer (500 mM Tris-HCl, pH 7.5; 50 mM MgCl2; 10 mM DTT), 3 μl of dNTP mix (10 mM), 1 μl of RNase H (2U/μl), 5 μl of DNA Pol I (10U/μl) and 61 μl water. The mixtures were incubated for 2 h at 16°C. The libraries were constructed with Nextera DNA Sample Preparation Kit (Illumina) per the manufacturer’s protocol. Generally, double-strand cDNA was fragmented, subjected to adapter ligation and amplified.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Data processing |
Illumina HiSeq 2500 sequencing, 50bp reads.
Adapter removal with cutadapt v1.16
Alignment to Mus musculus genome with hisat v2.1.0
Assignment of reads to gene features with Subread featureCounts v1.6.1
Signal detection via differential expression analysis between triplicate groups with DESeq2 1.18.1
Genome_build: mm10
Supplementary_files_format_and_content: Number of reads assigned to gene features, as computed by Subread featureCounts v1.6.1
|
| |
|
| Submission date |
Jul 26, 2018 |
| Last update date |
Oct 23, 2019 |
| Contact name |
Cameron Palmer |
| E-mail(s) |
palmercd@nih.gov
|
| Organization name |
National Institutes of Health
|
| Department |
National Institute of Diabetes and Digestive and Kidney Diseases
|
| Lab |
Laboratory of Cellular and Developmental Biology
|
| Street address |
50 South Drive
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL17021 |
| Series (2) |
| GSE117706 |
‘Sertoli cell-only’ phenotype and single-cell RNA-seq define PRAMEF12 as a germline-specific factor for male stem cell maintenance and differentiation [RNA-Seq] |
| GSE117708 |
‘Sertoli cell-only’ phenotype and single-cell RNA-seq define PRAMEF12 as a germline-specific factor for male stem cell maintenance and differentiation |
|
| Relations |
| BioSample |
SAMN09724203 |
| SRA |
SRX4472972 |
