title: Transcriptional profiling using a novel cDNA array identifies differential gene expression during porcine embryo elongation.
abstract: A novel porcine cDNA array, containing 1,015 PCR products selected for embryonic expression, was used for transcriptional profiling of conceptuses at four stages of peri-implantation development. Total conceptus RNA from small spherical, large spherical, tubular, and filamentous stages was amplified, converted to cDNA, and hybridized to membranes. Initially, normalized signal intensities obtained using cDNA from total RNA or from amplified RNA were compared. Uniform distribution of P-values associated with t-tests conducted for each gene indicated no evidence that amplification introduced bias. Analysis of data obtained by using amplified targets and the novel array identified genes differentially expressed across stages. Such genes were identified by testing for significant stage effects in gene-specific mixed models. A total of nine genes were declared differentially expressed. Six of the nine genes had P-values less than 0.001, and a false discovery rate of approximately 17% was associated with this significance threshold. Two out of six genes were significant when using the Bonferroni method to control the probability of one or more false positives. The other three genes had P-values between 0.001 and 0.01 and exhibited differences greater than twofold between stages. All four genes selected for confirmation (steroidogenic acute regulatory protein, interleukin 1 beta, transforming growth factor beta 3, and thymosin beta 10) were shown to be differentially expressed by using quantitative real time RT-PCR. Our study shows that RNA amplification is useful for transcriptional profiling with limiting porcine embryonic RNA, and that this novel targeted array can detect differential gene expression during trophoblastic elongation. Finally, our results contribute to an increased understanding of the temporal patterns of expression of known genes controlling conceptus development, as well as identify novel genes also differentially regulated during implantation.
authors.affiliation: Department of Animal Science, Iowa State University, Ames, Iowa 50011, USA.
authors.forename: So Hyun
authors.initials: SH
authors.lastname: Lee
authors.forename: Shu-Hong
authors.initials: SH
authors.lastname: Zhao
authors.forename: Justin C
authors.initials: JC
authors.lastname: Recknor
authors.forename: Dan
authors.initials: D
authors.lastname: Nettleton
authors.forename: Sarah
authors.initials: S
authors.lastname: Orley
authors.forename: Sung-Keun
authors.initials: SK
authors.lastname: Kang
authors.forename: Byeong-Chun
authors.initials: BC
authors.lastname: Lee
authors.forename: Woo-Suk
authors.initials: WS
authors.lastname: Hwang
authors.forename: Christopher K
authors.initials: CK
authors.lastname: Tuggle
mesh_terms: D000818:Animals; D020869:Gene Expression Profiling; D018507:Gene Expression Regulation, Developmental / Q000502:physiology*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D034421:Sus scrofa / Q000196:embryology*