Identification, Characteration Of Expression, SNPs Detection, Prokaryotic Expression And Association Anlysis Of The Eight Genes With Part Traits In Beef Cattle

Obtainning larger economic benefit is the final purpose in cattle breeding. So all the traits should be thought thoroughly, which may influence the economic benefit of cattle for a long time. Based on this idea, it is not only the major production traits, directly to the benefits, but also the secondary traits like productivity, lifespan, calf behavior and resistence of disease, which influences the benefits indirectly, should be taken attention on. For these traits are influenced by the conditions, heredity, and nutrition and so on, and difficult to measurement, the secondary traits selection plannings become so complecity and difficulty. And because the heritability of the secondary traits is very low, it results in slower progression of breeding with common methods. In order to provide certain methodology, what could it reveal the genetics roles of the secondary traits? This article focused on the molecule genetics of some lifespan traits genes, which influence the health and efficency, to provide elementary genetic information to cattle breeding by modern molecule and quantitative combination breeding methods, such as ESTs, RT-PCR, GenScan, direct sequencing, RFLP, Q-PCR, prokaryotic expression, bioinfermtics methods and association analysis. And the results were showed below:1. The complete CDS sequences of AKAP10,MBL2,PON1,P53,PON2,SIRT2,SIRT3 and WARS2 cDNA were cloned. And the cDNA sequences of AKAP10, PON1, PON2 and SIRT2 genes were submitted to GenBank, the accession numbers were EU239246, EU289337, EF522785 and EU562194, respectively.2. Five candidate genes (AKAP10,PON1,PON2,MBL2 and WARS2) were detected in ovary, liver, skeleton muscle, small intestine, fat, uterus, kidney, heart, lung, lymph, rumen, testis and galactophore by RT-PCR and Real-time PCR, and the results showed that 4 candidate genes are higher expression in liver except that AKAP10 gene is highly expressed in testis specially. Comparing with the database of electron expression in NCBI website, there were obviously differences between the database and the results.3. PON1 and PON2 genes'exons sequences were detected for SNPs and 6 SNPs were identified: A147G and G162A in exon6 of PON1 gene; A73G and A110G in exon5, T76C in exon6, T98C in exon9 of PON2 gene. Among these SNPs, the hereditary capacities of A147G, G162A in PON1 gene and T98C in PON2 gene were analyzed by PCR-RFLP and PCR-SSCP in 12 bovine breeds, respectively.4. Association analysis between PON1 and PON2 genes SNPs and body size, growth and parts carcass traits were carried out. (1) It is significant difference between body size and stem length, and chest measurement, and circumference of cannon bone and abdomen circumference (P<0.05), except for height at withers. (2) Individuals of AA genotype at A162G in exon6 of PON1 gene may act as short term cram group, and GG type should be eliminated through selection or competition. Individuals of GA types possess line fitting follow the ages, this type may have reasonable three-dimensional structure, and growth and development pattern in reproduction group. So A162G locus should be tasted to predict cattle lifespan. (3) The results of two enzymes digesting analysis shown that the specialized beef breed grow fast than dual purpose breeds. Simmental is in inferior position in the carcass traits, except for backfat thickness, tendness, total bone weight, rib eye area and carnation traits. At the same time, the beginning weight difference of any haplotype individuals maybe from some growth potential. The AeAeGaGa genotype of PON1 gene may influence the backfat thickness directly (P<0.01). (4) The results of association anlysis in 7 breeds shown that the rib eye area is higher in the individuals of BB genotype than those in AA and AB types significantly (P=0.0302). (5) There were significant differences of three genotypes in exon9 of PON2 gene among 7 breeds (P<0.01), and no difference in or between breeds and in various age groups (P>0.05). However, there are significant differences among three genotypes (P<0.01). And three genotype frequencies present a kind of reducing tendency following ages, but AA and BB genotype frequencies emerge supernumerary phenomenon about at 7 years old.5. The prokaryotic expression vector of PON1 and PON2 genes were structured using RT-PCR and pET28a+ vector, and the PON1 and PON2 and 6 His fusion proteins were expressed in the BL21 strain. These results will provid the bases for the purifying and application of these two fusion proteins.