Evaluation of candidate reference genes for investigation of the uterine tissue and corpus luteum of pigs on day 6 after artificial insemination

Abstract

Housekeeping genes (HKG) are genes necessary for the maintenance of basal cellular functions, regardless of the specific roles within a tissue. It, therefore, is expected that these genes will maintain a relatively constant expression profile when there are varying physiological conditions. The identification of tissue specific reference genes is highly important for the normalization of gene expression profiles among different tissues. In this sow study, the objective was to identify stable reference genes in the uterine tissue and corpus luteum (CL), 6 days post-artificial insemination. The stability of ubiquitin (UBB), glyceraldehyde 3-phosphate dehydrogenase (GAPDH), DNA topoisomerase 2-beta (TOP2B), histone H3 (H3F3A) and hypoxanthine-guanine phosphoribosyltransferase 1 (HPRT1) abundances of mRNA were evaluated using the Bestkeeper technique. Briefly, total RNA was extracted for each tissue from 20 gilts (n = 20), processed by RT-qPCR and submitted to analysis using the Bestkeeper technique, which allowed for the evaluation of the consistency in abundance of mRNA for the reference genes. For all evaluated genes, the abundance of mRNA was relatively consistent in the uterine tissue, with the greatest abundance being for the GAPDH and TOP2B genes. The analysis of these genes in the CL, however, indicated there was a relatively greater variation of mRNA abundance for the various reference genes. Data suggest that UBB was the reference gene with the most consistent relative abundance of mRNA and that this gene could be used as a reference for corpora lutea analyses of mRNA.