Development of Molecular Techniques to Better Identify the BoLA Class I Genes

Abstract

Innate resistance of a host to infection is strongly under the influence of genetic factors. Genes coding for histocompatibility antigens are a cluster of genes called the major histocompatibility complex (MHC). MHC gene products play a central role in the immune system. The MHC encodes families of class I, class II, and class III genes. The bovine lymphocyte antigen (BoLA) system is the major histocompatibility complex of cattle. BoLA class I genes are possibly involved in resistance to brucellosis in cattle. In this study, lymphocytes from animals which were previously grouped as resistant or susceptible phenotypically by conjunctival inoculation of 1x10⁷ CFU of live Brucella abortus were isolated over Histopaque-1077. mRNA was isolated from lymphocytes using the Micro-Fast track mRNA isolation kit from Invitrogen. Isolated mRNA was used to synthesize cDNA and this cDNA was employed in PCR reactions using BoLA class I gene specific primers. PCR products were analyzed on polyacrylamide gels. These PCR products were cloned and sequenced using Sanger Dideoxy method. In this investigation, molecular biological techniques were developed to better identify the major histocompatibility complex Class I genes in cattle and possible co-segregation of Class I genes with resistance or susceptibility to brucellosis was studied.