|
| Status |
Public on Sep 10, 2016 |
| Title |
Scr 2 |
| Sample type |
SRA |
| |
|
| Source name |
Control oocytes
|
| Organism |
Mus musculus |
| Characteristics |
strain: (C57BL x CBA) F1
developmental stage: Metaphase II
|
| Treatment protocol |
injections with siRNAs
|
| Growth protocol |
Ovaries were dissected from two to five 10-12 day old (C57BL x CBA) F1 females. To obtain individual follicles, the ovaries were incubated in modified MEM-alpha medium optimized for in vitro culture of follicles (MEM-alpha (Gibco 12000-014) supplemented with 0.026M NaHCO3 (Sigma), 5678U/100 ml Penicillin G (Sigma) and 8265U/100ml Streptomycin (Sigma), 1x ITS (Insulin/Transferrin/Selenium Solution; Sigma; Stock is 100x), 5% FBS (Fetal Bovine Serum; Gibco 16000044) and 0.01 µg/ml FSH (Follicle Stimulating Hormone; National Hormone and Peptide Program, NDDK-oFSH-20) that was supplemented with 2 mg/ml collagenase (Roche) for about 30-40 minutes total. During incubation with collagenase, the ovaries were pipetted up and down every 10 minutes to facilitate dissociation and then washed through several droplets of follicle culture medium without collagenase and were cultured at 37°C in 5% CO2 on membrane inserts in 6 well or 12 well culture dishes filled with follicle culture medium (see above). After 8-10 days of culture the oocytes were isolated in M2 medium supplemented with 10% FBS and matured until reached metaphase of the second meiotic division (MII).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
NucleoSpin RNA XS (Macherey-Nagel)
A cDNA library was prepared using SMARTer® Ultra™Low Input RNA for Sequencing (Clonetech Laboratories) and the samples were processed by BGI Tech Solutions. The cDNA product was synthesized and amplified using SMARTer PCR cDNA Synthesis Kit (Clontech) from the total RNA (10 ng) of sample. The cDNA was fragment by Covaris E210 (Covaris) and the median insert length was about 200bp.
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
| |
|
| Data processing |
Illumina OLB(off-line Basecaller) software used for basecalling.
Sequenced reads were trimmed for adaptor sequence, and masked for low-complexity or low-quality sequence, then mapped to mm9 whole genome using bwa with parameters GenomeBwaOptions_PE = -o 1 -e 63 -i 90 -L -k 2 -l 31 -t 4 -q 10
The mapped read counts for each gene were normalized for RNA length and for the total read number in each lane using the fragments per kilobase per million (FPKM) method.We used a strict algorithm to identify differentially expressed genes (DEGs) based on comparing the exposed with unexposed group to generate the DEGs.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited excel files include FPKM values for each Sample
|
| |
|
| Submission date |
Jun 29, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Michał Pasternak |
| E-mail(s) |
mpasternak.student@gmail.com
|
| Phone |
+447895431334
|
| Organization name |
MRC Laboratory of Molecular Biology
|
| Street address |
Francis Crick Avenue
|
| City |
Cambridge |
| State/province |
- None - |
| ZIP/Postal code |
CB2 0QH |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL21103 |
| Series (1) |
| GSE83876 |
Transcriptome comparison of oocytes treated with RNAi against Btg4 or with control siRNAs |
|
| Relations |
| BioSample |
SAMN05326602 |
| SRA |
SRX1885922 |
