Cloning and sequencing of cagA gene fragment of Helicobacter pylori with coccoid form

摘要

AIM: To clone and sequence the cagA gene fragment of Helicobacter pylon (H pylori) with coccoid form. METHODS: H pylori strain NCTC11637 were transformed to coccoid form by exposure to antibiotics in subinhibitory concentrations. The coccoid H pylori was collected. cagA gene of the coccoid H pylori strain was amplified by PCR. After purified, the target fragment was cloned into plasmid pMD-18T. The recombinant plasmid pMD-18T-cagA was transformed into E.coli JM109. Positive clones were screened and identified by PCR and digestion with restriction endonucleases. The sequence of inserted fragment was then analysed. RESULTS: cagA gene of 3 444 bp was obtained from the coccoid H pylori genome DNA. The recombinant plasmid pMD-18T-cagA was constructed, then it was digested by BamH Ⅰ+Sac Ⅰ, and the product of digestion was identical with the predicted one. Sequence analysis showed that the homology of coccoid and the reported original sequence H pylori was 99.7%. CONCLUSION: The recombinant plasmid containing cagA gene from coccoid H pylori has been constructed successfully. The coccoid H pylori contain completed cagA gene, which may be related to pathogenicity of them.