Preliminary Study Of Deer Tuberculosis Wild Strain And The BCG Discrepant Gene Prokaryotic Expression And Reactionogenicity Of Expression Products

Deer tuberculosis, which mainly caused by the Mycobacterium bovis, is common all over the world. It is a chronic, and analosis disease that is communicable between both live stocks and human. The pathology indicators are that Deer tuberculosis can form tuberculose focus, granuloma, and cheesy necrosis in organs. Mycobacterium can infect almost all kinds of deer, no matter wild or naturalized. It is true that deer tuberculosis or mycobacterium has been detected from almost all the countries. Since the first found of deer tuberculosis in New Zealand, Switzerland, America, Denmark, Canada, and Britain has all reported the infected deer with Deer tuberculosis. Ever after the discovery of Deer tuberculosis from deer farm in the northeast of China, the Deer tuberculosis has been the long lasting threat to Chinese deer farming. In the recent years, Deer tuberculosis has increasingly spreadable, as the popularity of human tuberculosis has increased, and also the host in wild animals, drug fast MTB, which incur a tremendous loss for Chinese deer industry.So far, there are still vacancies in the precaution and treatment area of Deer tuberculosis both abroad and at home, and the most common policy adopted by developed country is quarantine-catch and kill. In consideration of the high medical utility of deer, however, Chinese veterinarian use BCG to prevent deer tuberculosis. BCG is an attenuated vaccine of bovine mycobacterium. And BCG itself has some defects, such as the immune effects of instability in different parts of the body after vaccination. At the same time,BCG vaccination would interfere with PPD allergy testomg.Because of that, it is difficult to distinguish between vaccine immune animal and natural infection anmimal,vaccune strain and wild strain.The stuay show that there are16difference areas between M.tuberculosis H37Rv and BCG,named of RD1～RD9(totally contains61develop reading frame).The RD1area only exists in pathogenic mycobacteria but deletion in BCG.A long time,RDl area is considered to be the main virulence genes of Mycobacterium tuberculosis,the gene region encoding protein has a strong immunogenicity.As a result, we pick out some different genes from RD1area which have specific antigen and purified. We use SOE to obtain the unite antigen by different genes’fusion, in order to improve the serological diagnosis of deer tuberculosis have become prevailing studies on the world scene.In this study,we subcloned and sequenced DTB-2、 DTB-7、 DTB-46、 DTB-35which in the RD1area.We also make the gene fusion of DTB-46、 DTB-35, and analysis the reactogenicity by SDS-PAGE and Western bolt. Studies have shown that, DTB-2、 DTB-7、 DTB-46、 DTB-35are the major virulence genes of MTB as they have strong reactogenicity.So they can be used as the candidate antigen of the serum diagnosis of Deer tuberculosis.The existing studies have shown that the gene fusion to be a coalition antigen have more sensitive than a single gene. First,we design some specific primer to amplification purpose gene by PCR, then link the purpose gene to PMD-18-T simple and sequencing。 And next, we link recombinant clonep plasmid to PGEX-4T-1to gain prokaryotic expression;at last, we analyze expression products’reactionogenicity by SDS-PAGE and Western bolt.In this research we make different genes’fusion to get the purified and specific unite antigen.In order to improve the sensitivity of serological diagnosis of deer tuberculosis, which solidify the foundation of the diagnosis and universal precaution of deer tuberculosis. It is meaningful to the deer farming in the long run.