Polymorphism Analysis Of MHC-DRB,DQB Genes PCR-RFLP In Dolang Sheep And Chinese Merino Sheep

Major Histocompatibility Complex ( MHC ) is a chromosomal region consisting of a group of closely linked loci that are highly polymorphic, which is generally found in almost all vertebrates. MHC gene products are a type of cell surface glycoproteins, involved in T and B lymphocyte-mediated immune reactions, and composed of an alpha and a beta chain. MHC plays a central role in immune system in vertebrates. MHC genes are extensively investigated as candidate genes for disease resistance in many domestic animal species, which is a hot topic in molecule immunogenetics.MHC of sheep ( i.e Ovine Lymphocyte Antigen,OLA ) was mapped to chromosome 20 and composed of class I, II and III molecules. Polymorphisms of OLA genes are focused on DR and DQ subareas of class II molecule that plays a central role in immune system. There are many genes of OLA class II molecules, but the genes only at the DR and DQ subareas are expressed. The second exon of the DRB and DQB genes have been widely investigated because of its extensive polymorphism. The polymorphisms of the second exon of OLA-DRB1 and OLA-DQB genes were investigated using PCR-RFLP method in Dolang sheep and Chinese Merino sheep, which provides a basis for further resistance breeding.The polymorphisms of the second exon of OLA-DRB1 gene were investigated using the nested PCR-RFLP method in Dolang sheep and Chinese Merino sheep. The results show that there are extensive polymorphisms at position SacI, Hin1I and HaeIII in the second exon of DRB1 gene in Dolang sheep and Chinese Merino sheep, and the loci were controlled by 2, 2 and 6 kinds of common dominant alleles, repectively. The frequency of allele HaeIII E in Dolang sheep was higher than that in Chinese Merino sheep, and frequency difference of HaeIII E was significant ( p<0.05 ) between Dolang sheep and Chinese Merino sheep, others alleles frequencies were not significant ( p>0.05 ). Twenty-five PCR-RFLP defined genotypes were found in two sheep breeds. The distribution of genotypes appeared no unanimous in two sheep breeds, twenty-four PCR-RFLP defined genotypes were found in Dolang sheep, and twenty-one in Chinese Merino sheep. The frequencies of SacⅠAB,Hin1ⅠAB,HaeⅢAB,HaeⅢAF HaeⅢCF and HaeⅢBE were higher in Dolang sheep than that in Chinese Merino sheep, and the frequencies of HaeⅢAC,HaeⅢCD and HaeⅢAA were higher in Chinese Merino sheep than that in Dolang sheep,difference of above genotypes frequencies were significant (p<0.01,p<0.05) between Dolang sheep and Chinese Merino sheep,others were not significan(tp>0.05). Keeping Hard-weinberg equilibrium plays an important role in maintaining the stability of colony genetic structure. It showed difference for Hardy-weinberg equilibrium of each OLA-DRB1 gene digested with endonucleases among sheep varieties. All sheep's HaeⅢdigestion of genomic site doesn't still attain Hardy-weinberg equilibrium ( p<0.01 ).Chinese Merino sheep obtain Hardy-weinberg equilibrium in SacI and Hin1I digestion of genomic site ( p>0.05 ). Dolang sheep doesn't still obtain Hardy-weinberg equilibrium in SacI and Hin1I digestion of genomic site (p<0.01 ).The polymorphisms of the second exon of OLA-DQB gene were investigated using PCR-RFLP technique in Dolang sheep and Chinese Merino sheep. The results show that there are extensive polymorphisms at position TaqI, HaeIII and MvaI in the second exon of DQB gene in Dolang sheep and Chinese Merino sheep, and the loci were controlled by 3, 3 and 6 kinds of common dominant alleles, repectively. According results of cloning and sequencing in this experiment, allele HaeⅢm,HaeⅢn,MvaⅠz and MvaⅠy are new alleles found in Dolang sheep and Chinese Merino sheep. The frequency of MvaⅠc and MvaⅠd in Dolang sheep was higher than that in Chinese Merino sheep, the frequency of MvaⅠy in Chinese Merino sheep was higher than that in Dolang sheep, and frequency difference of MvaⅠc, MvaⅠd and MvaⅠy was significant ( p<0.05, p<0.01 ) between Dolang sheep and Chinese Merino sheep, others were not significant ( p>0.05 ). Twenty-six PCR-RFLP defined genotypes were found in two sheep breeds. The distribution of genotypes appeared no unanimous in two sheep breeds, twenty-one PCR-RFLP defined genotypes were found in Dolang sheep, and twenty-six in Chinese Merino sheep. The frequency of MvaⅠYY was higher in Chinese Merino sheep than that in Dolang sheep, and the frequency of MvaⅠCD was higher in Dolang sheep than that in Chinese Merino sheep, frequency difference of MvaⅠYY and MvaⅠCD were significant (p<0.01) between Dolang sheep and Chinese Merino sheep,others were not significant(p>0.05). It showed difference for Hardy-weinberg equilibrium of each OLA-DQB gene digested with endonucleases among sheep varieties. The results ofχ2 test shows that all sheep's TaqI digestion of genomic site attain Hardy-weinberg equilibrium ( p>0.05 ).Chinese Merino sheep and Dolang sheep doesn't still obtain Hardy-weinberg equilibrium in HaeIII and MvaI digestion of genomic site (p<0.01 ) .