Comparative Gene Expression Profiling of In Vivo-Derived Expanded and Hatched Porcine Blastocyst-Stage Embryos.

Abstract

During the pre-implantation period of embryonic development, the mammalian embryo exhibits dramatic changes and many key events take place. The developmental stage at which porcine embryos are most commonly manipulated is the blastocyst, and the quality of the embryos used is critical to the success of reproductive technologies such as embryo transfer. Despite the importance of this critical period, our ability to determine early embryonic quality based on morphological criteria is limited. As an alternative, characterization of the gene expression profile of the early porcine embryo could identify gene markers of embryonic quality. The objective of this study was to perform comparative gene expression profiling analysis of in vivo-derived expanded and hatched porcine blastocysts using two-colored microarray (Pig-Oligo Array, USDA Pig Genome Consortium). Microarray data were interpreted using WebArray and DAVID functional annotation tools. The microarray results identified 1783 genes that were commonly expressed in both expanded and hatched porcine blastocysts. A further 148 genes were differentially expressed (p<0.05) and in comparison to expanded blastocysts, 35 genes were down-regulated and 113 genes were up-regulated after hatching. Many of the differentially expressed genes are related to important molecular mechanisms, such as macromolecule metabolism and energy related pathways. A total of 14 genes of interest (LDHA, LDHB, SLC16A7 (previously known as MCT2), BSG, SLC2A1, SLC2A3, SLC2A5, POU5F1 (previously known as OCT4), ACTB, GAPDH, GRB2, SETX, SPG7, and YWHAZ) were selected from the gene list generated by the microarray analysis, and their expressions were verified using SYBR Green-based real-time PCR. For the genes tested, the real-time PCR results were consistent with the microarray results. The differences detected in the gene expression profiles between in vivo-derived expanded and hatched porcine blastocysts increases our understanding of the molecular mechanisms involved in the hatching process and identifies potential candidate gene markers of porcine embryo quality. This research was supported by the Natural Sciences and Engineering Research Council (NSERC) and is a part of the activities of the EmbryoGENE NSERC Strategic Research Network. (poster)