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Comparison of small medium and large healthy ovarian pig follicles

ABSTRACT: Three different stages of pig antral follicles have been studied in a granulosa-cell transcriptome analysis on nylon microarrays (1152 clones). The data have been generated from 7 RNA follicle pools and several technical replicates were made. Four Large, one Medium and two Small follicles pools were considered. For each follicle pool, 2 radioactive labellings were performed. Each membrane was exposed 16 hours (to avoid saturation of the signal of highly expressed genes) and 28 hours (to get some signal from lowly expressed genes). Each probe was hybridised on GPL3971 scag_scai Sus scrofa 1.2K mono array and on GPL3970 scag_scai Sus scrofa 4.6K triplicate array (except for GFS2172 which was labelled only once and hybridised onto 2 GPL3971 scag_scai Sus scrofa 1.2K mono array membranes), so that 4 spots are available for each gene (and 2 spots for GFS2172), for a given RNA and a given radioactive labelling. Keywords: granulosa-cell transcriptome analysis

ORGANISM(S): Sus scrofa

PROVIDER: GSE5299 | GEO | 2008/06/12

SECONDARY ACCESSION(S): PRJNA95399

REPOSITORIES: GEO

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Project description:The aim of this study is the identification of genes and gene networks involved in pig ovarian follicular development. cDNA nylon micro-arrays (2849 sequenced clones) were designed from different libraries : four subtractive suppressive hybridization libraries (generated from small versus large and small versus medium follicles) and a pig multi-tissue cDNA library. Granulosa cells were isolated from healthy follicles (small, medium or large), 24 h or 96 h after the end of a progestagen treatment. The RNA isolated from these cells was used to hybridize the micro-arrays and hybridisations were performed in duplicate. The images were quantified using Bzscan software and the data were managed with BASE software. Statistical analysis was performed using R software. Keywords: cell type comparison Granulosa cells were isolated from healthy follicles (small, medium or large). The RNA isolated from these cells was used to hybridize nylon micro-arrays. The arrays contained PCR products from 2848 pig cDNA clones (GPL3978), spotted in duplicate on two separate fields of the same membrane (last number 1 of the local ID corresponds to the left field and 2 corresponds to right field). The data have been generated from 14 RNA follicle pools (five Large, five Medium and four Small follicles pools). For each follicle pool, 2 radioactive labellings were performed (before last number of the local ID corresponds to the duplicate number). Each membrane was exposed 6, 12 or 24 hours (depending on the saturation signal) to radioisotopic-sensitive imaging plates (BAS-2025, Fujifilm, Raytest France S.A.R.L., Courbevoie, France). The imaging plates were scanned thereafter with a phosphor imaging system at a 25 µm resolution (BAS-5000, Fujifilm, Raytest France S.A.R.L., Courbevoie, France). The images were quantified using semi-automated software, BZScan (Lopez F, Rougemont J, Loriod B, Bourgeois A, Loi L, Bertucci F, Hingamp P, Houlgatte R, and Granjeaud S. Feature extraction and signal processing for nylon DNA microarrays. BMC Genomics 5: 38, 2004). Fixed circle segmentation, i.e. a grid process with a fixed spot diameter was applied. The hybridization images were quantified by the extraction of the intensity for each spot. A new version of BZsan (AGscan solfware) is available at : http://mulcyber.toulouse.inra.fr/projects/agscan/ The experimental data were managed using the free BASE software (Saal LH, Troein C, Vallon-Christersson J, Gruvberger S, Borg A, and Peterson C. BioArray Software Environment (BASE): a platform for comprehensive management and analysis of microarray data. Genome Biol 3: SOFTWARE0003, 2002.) modified by Sigenae team to accept radioactive experiments. Finally, data consisted in (14 RNA x 2 labellings x 2 fields) 14 probes, 28 hybridisations, 42 images (14 images were filtred out for background too hight).

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Comparison of small medium and large healthy ovarian pig follicles

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