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Sample GSM572678 Query DataSets for GSM572678
Status Public on Dec 31, 2012
Title Bt-2344-180-26-0308-7j
Sample type RNA
 
Source name Total RNA extracted from a pool of 15 to 20 mm antral follicles, determined as healthy using microscopic examination after Feulgen staining of some granulosa cells, originating from cow 400
Organism Bos taurus
Characteristics annotation pf/mf/gf: GF
group: large follicles
date hybridation-genovul: 2008-03-11
status: S (healthy follicles)
animal-number_genovul: 400
Extracted molecule total RNA
Extraction protocol Once ovaries collected, all visible antral follicles larger than 1 mm in diameter for sows and 3 mm for cows were isolated carefully using a binocular microscope. After dissection, follicle diameter was measured and each follicle was classified according to size, animal and follicular quality. Follicles were allocated to two size classes: small (noted SF, 1-2 mm for sows and 3-5mm for cows) and large (noted LF, 7-8 mm for sows and 15-20mm for cows). Granulosa cells were recovered from all individual follicles and they were stored at -80°C until RNA extraction. For each follicle, a sample of granulosa cells was smeared on a histological slide, and then Feulgen stained to determine follicular quality by microscopic examination. Only healthy follicles (presence of mitosis and absence of pycnosis in granulosa cells) were kept for this study. RNA was extracted from granulosa cells according to the technique described by Chomczynski and Sacchi with minor modifications using pools of granulosa cells from the same follicle size class and the same animal. The quality of each RNA sample was checked through the Bioanalyser Agilent 2100 (Agilent Technologies, Massy, France) and low-quality RNA preparations were discarded.
Label P33
Label protocol The arrays were hybridized with 33P-labelled complex probes synthesized from 5 µg of each RNA sample, using SuperScript II RNAse H-reverse transcriptase (Invitrogen, Cergy-Pontoise, France)
 
Hybridization protocol Micro-arrays were first hybridized with a 33P-labelled oligonucleotide sequence present in all PCR products to control the quality of the spotting process and to evaluate the amount of DNA in each spot. After stripping, the arrays were hybridized with 33P-labelled complex probes synthesized from 5 µg of each RNA sample, using SuperScript II RNAse H-reverse transcriptase (Invitrogen, Cergy-Pontoise, France). Each complex probe has been hybridized on membrane exposed 1, 3 or 7 days to radioisotopic-sensitive imaging plates (BAS-2025, Fujifilm, Raytest, Courbevoie, France).
Scan protocol The imaging plates were scanned thereafter with a phosphor imaging system at 25µm resolution (BAS-5000, Fujifilm, Raytest, Courbevoie, France). Hybridization images obtained from oligonucleotide and complex probes were quantified using the semi-automated AGScan software.
Description A pig nylon micro-array is hybridized with a complex 33P labelled probe
Data processing Data exploration and analysis were performed using R software. The data coming from complex probes hybridization were analysed on the logarithmic scale. As a first step, negative spot with >7 intensity values were checked on images and replaced by the median of negative spots in case of obvious overshining effect or hybridization stain. Then, luciferase positive controls were removed and the correlations of the data from each membrane were examined intra condition. Hybridizations with a lower than 0.80 correlation coefficient were discarded. Spots with low signal value (below the average of empty spots + 3 standard deviations) were considered as unexpressed and were excluded from the analysis. Finally, the negative control spots were removed and the remaining data were centred for each membrane.
 
Submission date Jul 30, 2010
Last update date Dec 31, 2012
Contact name Gwenola Tosser-Klopp
E-mail(s) gwenola.tosser@toulouse.inra.fr
Phone 33 5 61 28 51 14
Organization name INRA
Lab Genetique Cellulaire
Street address Chemin de Borde-Rouge BP 52627
City Castanet-Tolosan Cedex
ZIP/Postal code 31326
Country France
 
Platform ID GPL3729
Series (2)
GSE23335 SABRE-Bt-Ssc_020410
GSE24221 Genovul-Bt-Ssc_020410

Data table header descriptions
ID_REF
VALUE see data processing protocol
QIM_CONST image signal intensity constant diameter
QFIT_VAR fit signal intensity varariable diameter
FIT_CORR fot correction
QIM_VAR image signal intensity variable diameter
QFIT_CONST fit signal intensity constant diameter
DIAM variable diameter
SPOT_QUAL spot quality
OVER_CORR overshining correction
QM quality measure

Data table
ID_REF VALUE QIM_CONST QFIT_VAR FIT_CORR QIM_VAR QFIT_CONST DIAM SPOT_QUAL OVER_CORR QM
CVU03687 null 366 427 -0.00018 427 365 341.697 1 0 0.83394
scan0030.h.10 -0.384073918 126 146 0 158 115 350 0 0 0.86341
scan0024.i.17 null 110 134 0 144 107 350 0 0 0.82486
scan0018.i.20 null 74 92 0 99 73 350 0 0 0.80385
scan0012.e.11 -0.345158502 131 163 0.00223 165 131 350 0 0 0.86686
scan0006.g.08 -0.484157377 114 88 0 155 71 350 0 0 0.70043
scaj0002.k.04 1.202891138 616 651 -0.00109 648 617 315.523 1 0 0.86413
scac0041.n.08 -0.656007634 96 64 0 138 51 350 0 0 0.60961
scac0033.p.05 null 110 142 0.00331 142 111 350 0 0 0.86174
scac0029.i.06 -0.731719455 89 60 0 114 48 350 0 0 0.67644
scac0025.g.14 null 93 62 0 126 49 350 0 0 0.64981
CVU03687_1 null 197 253 0.00446 258 202 350 0 0 0.86317
scan0036.n.11 null 125 28 0.00551 29 128 115.989 0 0 0.8585
scan0030.h.17 0.016086138 188 186 0 229 151 350 0 0 0.80144
scan0024.i.20 null 99 82 0 130 67 350 0 0 0.75206
scan0018.i.22 null 96 77 0 124 61 350 0 0 0.73408
scan0012.e.18 null 144 89 0.00509 90 146 216.757 0 0 0.87558
scan0006.g.17 -0.368325561 128 37 0.00568 37 131 155.839 0 0 0.9054
scaj0002.p.11 0.466619531 295 201 0.00684 203 302 219.287 0 0 0.90865
scac0041.n.11 -0.566395475 105 10 0.00465 10 105 82.4088 0 0 0.94734

Total number of rows: 9216

Table truncated, full table size 554 Kbytes.




Supplementary data files not provided
Processed data included within Sample table

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