Perfil transcriptômico de oócitos maturados e blastocistos bubalinos (Bubalus bubalis) produzidos in vitro.

Abstract

The buffalos have economic and industrial potential, which is, however, underutilized due, among other factors, to the low efficiency of reproductive biotechnologies such as In Vitro Embryo Production (PIVE). The efficiency of PIVE, in turn, is related to knowledge about the functioning of oocytes and embryos. In this sense, the use of New Generation Sequencing (NGS) of RNA (RNA-seq) has provided advances for the understanding of the mechanisms of oocyte maturation and embryo development in several domestic species, and were reviewed in chapter 1. This work is dedicated To study the molecular aspects of oocyte maturation and embryonic development in buffalos. In chapter 2, the description and analysis of the transcriptomes of mature oocytes and blastocysts produced in vitro of buffalos was made. For this, mRNA was extracted for the construction of barcoded libraries, which were sequenced on the Proton ™ Ion platform. The reads were aligned to the bovine reference genome (Bos taurus UMD3.1), using the Cufflinks program to calculate the relative abundance of the transcripts and the DESeq2 package to analyze expression. It was observed the expression of 13,976 genes, of which there were shared (62%) and exclusive genes with specialized functions in oocytes (1.6%) and blastocysts (15.7%). There were 4,153 genes differentially expressed in blastocysts in relation to oocytes, of which some genes with greater variation in expression were related to lipid metabolism, implantation and oocyte maturation. A transcriptomic panel of oocytes and in vitro blastocysts of buffalos was elaborated, and the importance of promising target genes was discussed in future strategies for enhancement of PIVE in the buffalo species. In chapter 3, transcripts of oocytes and buffalo blastocysts were compared to those of cattle reported in the literature. Since the PIVE protocols in buffalos are largely bovine inspired, this comparison was used to indicate the mimicry of the transcriptomes of these species. The analysis of coexpression networks and preservation of modules of the WGCNA package of the R program was used. As a result, a great similarity of buffalo and bovine transcriptomic profiles was observed. Of the 7 co-expression modules identified in buffalo, 4 were strongly preserved (Z-summary> 10) in cattle, with their genetic ontologies related to the embryonic development program. However, the expression profile within the modules, defined by the genes hub, was different, indicating important differences in terms of gene interactions between the two species. It is concluded that this initial analysis of oocyte and buffalo blastocyst transcripts indicated the presence of genes relevant for oocyte and embryonic quality. And the comparison of buffalo and bovine transcriptomes indicated differences of expression within conserved modules that justify the elaboration of specific PIVE protocols for buffalos.