|
| Status |
Public on May 07, 2012 |
| Title |
GV oocyte_Ring1/Rnf2 double mut_rep3 |
| Sample type |
RNA |
| |
|
| Source name |
GV oocyte, Ring1/Rnf2 double mutant
|
| Organism |
Mus musculus |
| Characteristics |
tissue: GV oocyte
strain: mixed 129/Sv and C57BL/6J
genotype/variation: Ring1-/-Rnf2F/FZp3-cre (Ring1/Rnf2 double mutant)
|
| Growth protocol |
Mouse oocytes were derived from superovulated 5-10 week old females according to standard procedures. Fully-grown germinal vesicle (GV)-intact oocytes were collected 46 h after PMSG injection (5 U, Intervet) in M2 medium (Sigma) containing 2.5 μM milrinone (Sigma).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
GV oocytes were pooled from several mice and RNA was isolated from batches of 50 oocytes, 3 biological replicates per genotype. RNA was isolated using the PicoPureTM RNA Isolation Kit (KIT0202) according to the manufacturer’s instructions (Stratagene). The quality of the RNA was assessed using the Agilent 2100 bioanalyzer and RNA 6000 Pico Chip. The extracted RNA was converted into OmniPlex WTA cDNA libraries and amplified by WTA PCR using reagents supplied with the TransPlex Whole Transcriptome Amplification kit (WTA1, Sigma, USA) following the manufacturer’s instructions with minor modifications. The obtained cDNA was purified using the GeneChip cDNA Sample Cleanup Module (Affymetrix).
|
| Label |
biotin
|
| Label protocol |
RNA was processed according to the GeneChip Whole Transcript (WT) Double-Stranded Target Assay Manual from Affymetrix (GeneChip Whole Transcription Sense Target Labeling technical manual, Rev. 2).
|
| |
|
| Hybridization protocol |
Affymetrix GeneChip arrays were hybridized following the GeneChip Whole Transcript (WT) Sense Target Labeling Assay Manual (Affymetrix) with a hybridization time of 16h. The Affymetrix Fluidics protocol FS450_0007 was used for washing.
|
| Scan protocol |
Scanning was performed with Affymetrix GCC Scan Control v. 3.0.0.1214 on a GeneChip® Scanner 3000 with autoloader.
|
| Data processing |
Probesets were summarized and probeset-level values normalized with justRMA() function from R (version 2.10.0) / Bioconductor (version 2.5) package affy using the CDF environment MoGene-1_0-st-v1.r3.cdf (as provided by Bioconductor) and annotation from Netaffx (www.netaffx.com) (provided as a supplementary file on the Series record).
|
| |
|
| Submission date |
Jul 20, 2010 |
| Last update date |
May 08, 2012 |
| Contact name |
Antoine Peters |
| E-mail(s) |
antoine.peters@fmi.ch
|
| Organization name |
Friedrich Miescher Institute for Biomedical Research (FMI)
|
| Street address |
Maulbeerstrasse 66
|
| City |
Basel |
| ZIP/Postal code |
4058 |
| Country |
Switzerland |
| |
|
| Platform ID |
GPL6246 |
| Series (2) |
| GSE23033 |
Polycomb function during oogenesis is required for mouse early embryonic development (germinal vesicle oocytes) |
| GSE28711 |
Polycomb function during oogenesis is required for mouse early embryonic development |
|
