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Sample GSM132658 Query DataSets for GSM132658
Status Public on Sep 05, 2006
Title Fully grown GV-stage oocyte sample #3
Sample type RNA
 
Source name oocyte
Organism Mus musculus
Characteristics GV-stage oocytes isolated from eCG-primed (44-46h) 22-24 days old female (C57BL/6JxSJL)F1 mice
Biomaterial provider The Jackson Laboratory
Treatment protocol Ovarian follicular development was stimulated by intraperitoneal injection of 5 IU of eCG (EMD Biosciences, Inc., Calbiochem, La Jolla, CA). Immature GV-stage oocytes were collected at 44-46 h post eCG priming by puncturing large antral follicles on ovarian surface with a pair of 26 gauge needles.
Growth protocol Released oocyte-cumulus cell complexes (OCCs) were collected and cumulus cells surrounding the oocyte were removed completely by passing OCCs several times through a hand-drawn small fine glass pipette with an inner diameter slightly narrower than the oocyte. Only those oocytes with an intact GV and no apparent sign of degeneration were collected. Both GV and MII oocytes were washed thoroughly in the collection medium to ensure that no cumulus cells were present. Medium used for oocyte collection was MEM-α (Invitrogen Corporation, Grand Island, NY) supplemented with 3 mg/ml of crystallized lyophilized bovine serum albumin (Sigma, St. Louis, MO), 75 mg/liter of penicillin G (Sigma) and 50 mg/liter streptomycin sulfate (Sigma). Milrinone (Sigma), a selective inhibitor of oocyte-specific phosphodiesterase (PDE3), was added into the medium at the concentration of 5 ?M to prevent the GV-stage oocytes from undergoing maturation.
Extracted molecule total RNA
Extraction protocol Total RNA was extracted from 300 oocytes using the Picopure RNA isolation Kit (Arcturus, Mountain View, CA).
Label Biotin
Label protocol Total RNA was extracted from oocytes using the PicoPure RNA Isolation Kit according to the manufacturer?s instruction. The RNA quality and yield of each sample were determined using the Bioanalyzer 2100 and RNA 6000 Pico LabChip assay (Agilent Technologies Inc, Palo Alto, CA) in combination with Quant-iTTM RiboGreen Reagent according to supplied protocols (Invitrogen, Carlsbad, CA). Ten nanograms of total RNA from each sample was employed in the two-round cDNA synthesis and subsequent in vitro-transcription according to the Two-Cycle Eukaryotic Target Labeling Assay (Affymetrix Expression Analysis Technical Manual: ? Section 2: Eukaryotic Sample and Array Processing? (http://www.affymetrix.com/support/technical/manual/expression_manual.affx)) with the following modifications. For the first and second cycle of cDNA synthesis steps, Full Spectrum? MultiStart Primers for in vitro transcription (SystemBiosciences, Mountain View, CA) were used, instead of Affymetrix?s standard T7-Oligo(dT) primer or random hexamers.
 
Hybridization protocol Standard AffyMetrix procedures
Scan protocol Standard AffyMetrix
Description Fully grown GV-stage oocytes isolated from 22-24d old eCG-primed mice.
Data processing Probe level data were imported into the R software environment and expression values were summarized using the RMA (Robust MultiChip Average) function (Irizarry et al., 2003) in the R/affy package with rma2 background adjustment, no inter-array normalization, and median polish summarization of perfect match values (Gautier et al., 2004). To account for the difference in starting RNA amount obtained from the same number of GV and MII oocytes, and based on the fact that there is no new transcriptional activity in the fully grown oocytes (De La Fuente, 2006), a normalization was performed on a sample of non-differentially expressed genes which were estimated and sampled using EM algorithm for a normal mixture model (Gautier et al., 2004; McLachlan and Krishnan, 1997)
 
Submission date Aug 29, 2006
Last update date Aug 28, 2018
Contact name You-Qiang Su
E-mail(s) youqiang.su@jax.org
Organization name The Jackson Laboratory
Street address 600 Main Street
City Bar Harbor
State/province ME
ZIP/Postal code 04609
Country USA
 
Platform ID GPL1261
Series (1)
GSE5668 Identification and characterization of the changed and stable transcripts during mouse oocyte maturation
Relations
Reanalyzed by GSE119085

Data table header descriptions
ID_REF
VALUE RMA

Data table
ID_REF VALUE
1415670_at 8.075974799
1415671_at 8.460421486
1415672_at 10.60062219
1415673_at 4.415702539
1415674_a_at 5.772777905
1415675_at 6.753307209
1415676_a_at 8.366023017
1415677_at 7.852756804
1415678_at 9.147688339
1415679_at 10.30382842
1415680_at 9.368170111
1415681_at 5.687043217
1415682_at 7.365628073
1415683_at 9.697319933
1415684_at 10.46587671
1415685_at 4.285159187
1415686_at 9.472640504
1415687_a_at 6.871417789
1415688_at 11.42090108
1415689_s_at 2.970895011

Total number of rows: 21702

Table truncated, full table size 497 Kbytes.




Supplementary file Size Download File type/resource
GSM132658.CEL.gz 5.9 Mb (ftp)(http) CEL
Raw data provided as supplementary file

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