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Transcriptional profiling by RNA-Seq of peri-attachment porcine embryos generated by a variety of assisted reproductive technologies.

Isom SC, Stevens JR, Li R, Spollen WG, Cox L, Spate LD, Murphy CN, Prather RS

Substantial mortality of in vitro manipulated porcine embryos is observed during peri-attachment development. Herein we describe our efforts to characterize the transcriptomes of embryonic disc (ED) and trophectoderm (TE) cells from porcine embryos derived from in vivo fertilization, in vitro fertilization (IVF), parthenogenetic oocyte activation (PA), and somatic cell nuclear transfer (SCNT) on days 10, 12, and 14 of gestation. The IVF, PA, and SCNT embryos were generated with in vitro matured oocytes and were cultured overnight in vitro before being transferred to recipient females. Sequencing of cDNA from the resulting embryonic samples was accomplished with the Genome Analyzer IIx platform from Illumina. Reads were aligned to a custom-built swine transcriptome. A generalized linear model was fit for ED and TE samples separately, accounting for embryo type, gestation day, and their interaction. Those genes with significant differences between embryo types were characterized in terms of gene ontologies and KEGG pathways. Transforming growth factor-β signaling was downregulated in the EDs of IVF embryos. In TE cells from IVF embryos, ubiquitin-mediated proteolysis and ErbB signaling were aberrantly regulated. Expression of genes involved in chromatin modification, gene silencing by RNA, and apoptosis was significantly disrupted in ED cells from SCNT embryos. In summary, we have used high-throughput sequencing technologies to compare gene expression profiles of various embryo types during peri-attachment development. We expect that these data will provide important insight into the root causes of (and possible opportunities for mitigation of) suboptimal development of embryos derived from assisted reproductive technologies.

DNA shearing

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July, 2013




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