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Sample GSM897432 Query DataSets for GSM897432
Status Public on Oct 14, 2012
Title Mus_adult_MII_4
Sample type RNA
 
Source name adult mouse follicles, mature in vivo, metaphase II, replicate 4
Organism Mus musculus
Characteristics tissue: adult ovarian follicles
Stage: mature_invivo
meiotic stage: metaphase II
development capacity: developmentally fully competent
Treatment protocol Cumulus cells were separated from the immature oocytes by gentle pipeting. In the case of expanded cumulus of ovulated COCs, they were transferred into M2-droplets containing 300 µg/ml of hyaluronidase (Sigma-Aldrich) for a few seconds to disperse granulosa cells. In all cases, media containing granulosa cells after oocyte denudation and elimination were centrifuged and cumulus cell pellets rinsed in PBS before snap-freezing in liquid nitrogen and storage at -70° until RNA extraction.
Growth protocol Immature, unexpanded cumulus-oocytes complexes (COC) were isolated from prepubertal (3-week-old) or adult (8-week-old) B6CBAF1/J mice ovaries by puncturing large follicles out of the ovaries in M2 culture medium (Sigma). Mature, expanded COCs were obtained from the oviduct from 8-week-old mice after standard superovulation protocol.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted using TRIzol reagent (Invitrogen, Cergy Pontoise, France). RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
Label Cy3
Label protocol Cyanine-3 (Cy3) labeled cRNA was prepared from 0.35 µg RNA using the One-Color Microarray based gene Expression kit (Quick Amp Labeling, Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
 
Hybridization protocol 1.65 µg of Cy3-labelled cRNA was fragmented at 60°C for 30 minutes in a reaction volume of 55 µl containing 25x fragmentation buffer and 10x blocking agent following the manufacturers instructions (Agilent Technologies, Santa Clara, CA) . On completion of the fragmentation reaction, 55 µl of 2x GEx Hybridization Buffer HI-RPM (Agilent provided) was added to the fragmentation mixture and hybridized to Agilent Bovine Oligo Microarray (G4846A) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent).
Scan protocol Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2566AA) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 5 µm, Dye channel is set to Green and Green PMT is set to 100%).
Description Gene expression in mouse ovulatory follicles in which oocytes have undergone maturation (metaphase II oocytes awaiting fertilization) and are developmentally fully competent.
Data processing The scanned images were analyzed with Feature Extraction Software 10.5.1.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid:026655_D_F_20100123) to obtain background subtracted and spatially detrended Processed Signal intensities. Poor quality features that were either saturated or non-uniform were flagged using Genespring Agilent software. Only features with a signal-to-noise ratio (SNR) of up to 2.6 and significantly different from the local background (two sided t-test) were used for further analysis. Entities were then filtered based on these flag values: at least 80% of the values in any 2 out of 3 conditions must have acceptable values. Expression values were then log2-transformed and submitted to a scale normalisation
 
Submission date Mar 19, 2012
Last update date Oct 14, 2012
Contact name Julien Bobe
E-mail(s) Julien.Bobe@rennes.inra.fr
Phone (33) 2 23 48 57 24
Organization name Institut National de la Recherche Agronomique
Lab INRA-SCRIBE
Street address Campus de Beaulieu
City Rennes
ZIP/Postal code 35000
Country France
 
Platform ID GPL11202
Series (2)
GSE36604 Transcriptional profiling of cumulus cells differentiation throughout oocyte competence acquisition in the murine ovarian follicles.
GSE36617 Contribution of molecular actors from somatic origin to oocyte developmental competence acquisition, lessons from evolution

Data table header descriptions
ID_REF
VALUE Normalized signal intensity (log2 values) calculated by Genespring Agilent GX10.0 software

Data table
ID_REF VALUE
GE_BrightCorner 13.927031
DarkCorner 3.834561
A_55_P1989846 8.854366
A_55_P2022211 16.290447
A_55_P1964375 11.303669
A_51_P128876 9.982538
A_51_P207591 9.387476
A_55_P2404223 9.112694
A_55_P2101944 16.117525
A_52_P358860 12.246426
A_51_P119031 9.573517
A_51_P309854 3.9343107
A_51_P343900 12.368696
A_51_P234359 7.464068
A_51_P487813 12.125074
A_52_P613977 12.091372
A_55_P2052210 14.876236
A_51_P128987 15.921234
A_55_P2111153 10.969759
A_51_P210560 13.027449

Total number of rows: 20880

Table truncated, full table size 471 Kbytes.




Supplementary file Size Download File type/resource
GSM897432_Mus_Adult_MII_4.txt.gz 2.2 Mb (ftp)(http) TXT
Processed data included within Sample table
Processed data are available on Series record

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