Cloning And Expression Of Helicobacter Pylori UreB And Its Immunoligical Activity Analysis

About fifty per cent of people in the world were infected by Helicobacter pylori (Hp) and Helicobacter pylori infection is being correlated to a number of human diseases.Urease is one of the most abundantly produced proteins by Hp.This enzyme can resolve urea into ammonia and carbon dioxide quickly.Anti-Urease antibody could be detected in the serum of patient infected by H. pylori.Now, based on urease,several methods have been established for diagnosing the presence of H. pylori infection.H. pylori cell is hard to be cultured,so gene recombination is developed to obtain a lot of protein of H. pylori. We express H. pylori ureB fragment to determine if it could be used as an antigen for serological test.H. pylori ureB fragments were amplified by PCR and subsequently cloned into the prokaryotic expression vector pBV220 after sequencing(The result showed that 97%of the DNA sequence we had got were the same as that of M60398,so they were the right gene we wanted).Then the DH5a was transformed by pBV220/ureB,and recombinant protein was induced by heat when temperature reached 42°C.the molecular mass of the expressed UreB were conformed to that we anticipated.The form of the recombinant UreB were inclusion bodies in host bacteria. Western-blot showed that the recombinant UreB was expressed in DH5a/ pBV220/ureB which could be recognized by anti-Hp antiserum .Then the recombinant UreB were used as the antigen of DIFGA to assay sera got from blood bank. The result is in agreement with that reported in the literature.For the higher purification efficiency,we also cloned the ureB into the prokaryotic expression vector pRSET-A, the recombinant UreB were expression after inducement,also were inclusion bodies.The protein were purificated with Ni-NTA affinity chromatograph,and SDS- polyacrylamine gel electro-phoresis showed single band .Used as antigen of DIFGA to assay sera got from blood bank, the result is in agreement with that reported in the literature.Our results indicated that the reactionogenicity of the recombinant UreB were not influenced by the diversity among Hp strains,and suggested that the recombinant UreB may be useful for serological test.For its non-invasive, fast, easy and inexpensive,it may be able to reach a reliable infection diagnosis. .