Expression analysis of rat testis at early development stage
ABSTRACT: Investigation of whole genome gene expression level changes in rat testes on the postnatal day 6, 8 and 10 to compare the differentially expressed genes between them. A three chip study using total RNA recovered from the testes of rat postnatal day 6, 8 and 10. There are 135,000 probes in a chip,each chip measures the expression level of 26419 genes of rat from Nimblegen. Fold-change screening between the two groups obtained from the experiment. The threshold used to screen up or down regulated genes is Fold change＞＝2.0
Project description:We performed RNA microarray in a low protein diet (LPD) model of IUGR at three key time points of alveolarization process. IUGR and control rat pups had been studied for each time point considered: on postnatal day 4 (P4) before beginning of the alveolarization process, on P10, peak of alveolarization process and on P21 at the end of it. A twelve chip study using total RNA extracted from 14 whole lungs intrauterine growth restricted rat pups compared to 15 controls. RNA was extracted at three key time points of alveolarization in postnatal life. RNA was pooled by groups of 2 ou 3 before analysis.
Project description:The purpose of this study is to assess the potential of TOTM to induce testicular maldevelopment in the rat. We studied the effects of TOTM on the expression of genes in pathways involved in steriodogenesis and testes development. Certain phthalate esters have previously been shown to be involved in the induction of rat testicular mal-development (TMD) through effects on the expression of genes in pathways involved in steroidogenesis and testes development. In order to assess the effects of a potential alternate plasticizer, tris(2-ethylhexyl)trimellitate (TOTM), rats were exposed daily, in utero, to TOTM in order to assess the potential of this compound to induce developmental effects on the fetal testes. Pregnant rats were exposed between gestational day 12 and 19 and fetal testes RNA was analysed using whole genome microarrays. The effects of TOTM on the expression of genes in pathways involved in steroidogenesis and testes development were examined. The effects of TOTM were also compared with di(2-ethylhexyl)phthalate (DEHP), mono(2-ethylhexyl)phthalate (MEHP), an active metabolite of DEHP, and 2-ethylhexanol (2-EH), which were used as positive and negative controls, respectively. MEHP & DEHP (500mg/kg) caused a repression of genes in TMD pathways involved in cholesterol synthesis and transport (HMGCS, HMGCR, StAR, SCARB1, FDFT1, FDPS), steroidogenesis (Cyp11a, HSD3B1, SC4MOL) and testes development (INSL3, INHA). 2-EH caused minor repression of some of the genes in the TMD pathway. This was rationalised on the basis that 2-EH, a DEHP metabolite, is also a weak PPARα agonist. It has been shown that in utero treatment with DBP will repress the genes from fetal testes involved in steroidogenesis and that this effect is associated with direct DBP-mediated binding of PPARα to the promoters of these genes (Plummer et. al. 2010). TOTM did not cause a significant repression of genes in the TMD pathway. Based on these data, it is highly unlikely that TOTM will cause testicular dysgenesis in rats. Rats were exposed to TOTM in utero by the administration of daily oral gavage doses to pregnant dams between gestational day 12 and 19. The foetal testes were obtained by micro-dissection and prepared to facilitate the isolation of RNA, which was subsequently analysed using whole rat genome microarrays.
Project description:The Drosophila spermatogenesis cell differentiation pathway involves the activation of a large set of genes in primary spermatocytes. Most of these genes are activated by testis-specific TATA-binding protein associated factors (tTAFs). In the current model for the activation mechanism, Polycomb plays a key role silencing these genes in the germline precursors, and tTAF-dependent activation in primary spermatocytes involves the displacement of Polycomb from gene promoters. We investigated the genome-wide binding of Polycomb in wild type and tTAF mutant testes. According to the model we expected to see a clear enhancement in Polycomb binding at tTAF-dependent spermatogenesis genes in tTAF mutant testes. However, we find little evidence for such an enhancement in tTAF mutant testes compared to wild type. To avoid problems arising from cellular heterogeneity in whole testis analysis, we further tested the model by analysing Polycomb binding in purified germline precursors, representing cells before tTAF-dependent gene activation. Although we find Polycomb associated with its canonical targets, we find little or no evidence of Polycomb at spermatogenesis genes. The lack of Polycomb at tTAF-dependent spermatogenesis genes in precursor cells argues against a model where Polycomb displacement is the mechanism of spermatogenesis gene activation. This genome-wide ChIP-array study investigates the binding of Polycomb in three biological samples: wild type (WT) whole testes, tTAF (can) mutant whole testes, and FACS-sorted germline precursor cells. We performed two biological replicates for each sample, except wild type whole testes where we performed three. For all ChIP-array experiments, input chromatin was used as the reference control to assay ChIP enrichment. We used Cy3/Cy5-labelled ChIP and input DNA for hybridisation onto Nimblegen arrays, and we performed a Cy3/Cy5 dye swap for one biological replicate of each sample (see supplementary file: GSE39935_README.txt).
Project description:High dose level dibutyl phthalate (DBP) exposure of fetal rat testes in vivo inhibits testosterone production (i.e. endocrine disruption). Here, fetal testis mRNA levels were profiled following exposure to a DBP dose level that did not significantly reduce testosterone levels. The goal was to identify the constellation of gene expression changes that do not correlate with endocrine disruption. Fischer 344 rats were exposed via oral gavage of the dam to vehicle (corn oil) or 50 mg/kg (body weight) DBP daily from gestational day (GD) 12 to 20. The day after mating was defined as gestational day 0. Six hours after the final exposure on GD20, fetal testes were dissected and mRNA levels quantified using Affymetrix Rat Expression 230 2.0 microarrays.
Project description:We conducted a whole transcriptome analysis of testes from a meiotic drive-carrying strain (T37) in comparison with a drive-sensitive strain (RED) using microarrays based on the complete annotated Ae. aegypti gene set. The T37 strain, which carries a strong meiotic drive gene (Mori et al., 2004 (PMID 15605641)), was established from mosquitoes collected in Trinidad. The RED strain is highly sensitive to the meiotic drive gene (Hickey and Craig, 1966 (PMID ); Mori et al., 2004 (PMID 15605641)). A six-chip study using total RNA recovered from three biological samples of the T37 strain and another three biological samples of the Red strain of Aedes aegypti. Each chip measures the expression level of 16,092 genes annotated from the Aedes aegypti genome sequence, with twenty 60-mer probe pairs (PM/MM) per gene, with three-fold technical redundancy.
Project description:The puropose of this experiment was to identify gene expression changes that result from 5-aza-2-deoxycytidine induced DNA demethylation of Swarm rat chondrosarcoma cells. The gene expression profiles of untreated Swarm rat chondrosarcoma cells were compared to the gene expression profiles of Swarm rat chondrosarcoma cells that were treated for 5 passages with a low dose of 5-aza-2-deoxycytidine (0.1uM). Five chip study. For these experiments, microarray was carried out on untreated (control) Swarm rat chondrosarcoma cells (3 biological replicates), and microarray was also carried out on Swarm rat chondrosarcoma cells treated with 5-aza-2-deoxycytidine (2 biological replicates).
Project description:The NuRD complex is required for efficient and timely myelination in the peripheral nervous system. ChIP-chip assays were performed on rat sciatic nerve at P15, a peak timepoint of myelination, for binding of Chd4 to genes involved in regulating myelin formation. This experiment includes two custom ChIP-chip design incorporating many genes that are dynamically regulated during myelination. The antibodies used in this platform were Chd3/4 (Santa Cruz sc-11378) Chd4 (gift from Paul Wade), Mta2 (Santa Cruz sc-9447), and Nab2 (Santa Cruz sc-22815). Chd4 ChIP samples from experimental and input samples were hybridized.
Project description:The Sertoli cells (Sc) of 5 days (infant) and 12 days (pubertal) old rat were isolated and cultured in triplicates. Nuclear and cytoplasmic fractionation was done for both the cases. All the four protein fractions (nuclear and cytoplasm of infant and pubertal Sc) were anaysed using Lc-MS/MS.
Project description:Egr2/Krox20 and Sox10 regulate genes involved in formation of myelination in the peripheral nervous system. ChIP-chip assays were performed on rat sciatic nerve at P15, a peak timepoint of myelination. In addition, Faire was used to identify areas of open chromatin. This experiment includes a custom ChIP-chip design incorporating many genes that are dynamically regulated during peripheral nerve myelination. Two antibodies were used for Egr2, Abcam and Covance PRB-236P. Egr2 and Sox10 ChIP samples were hybridized along with total input. In addition, FAIRE samples were hybridized relative to input DNA
Project description:ChIP microarrays were used to investigate whether dibutylphthalate (DBP)-mediated repression of SF1-regulated genes was associated with changes in transcription factor binding to genes involved in DBP-induced testicular maldevelopment. The repressive effect of DBP on SF1 regulated gene expression in fetal testes correlates with inhibition of SF1 binding to steroidogenic gene promoters. Comparison of control- and DBP- in utero 500mg/Kg treated fetal rat testes