幽门螺杆菌适应性定植相关基因HP0318的克隆及序列分析

目的 测定幽门螺杆菌(HP)适应性定植蒙古沙鼠前后HP0318基因序列并分析其序列差异, 以了解该基因在HP适应性定植过程中在基因水平是否发生了改变,为进一步研究HP的适应性变异机制提供实验依据.方法 对幽门螺杆菌临床分离株M0、沙鼠适应株M13及标准株NCTC11637基因组中的HP0318全长基因进行扩增,然后连接到PMD18-T载体进行测序,最后通过DNAsis软件对不同菌株来源的HP0318基因进行多重序列比对,并与Genbank公布的2个标准株(HP26695株和J99株)的HP0318序列进行对比分析.结果 PCR法成功扩增出了3个菌株的HP0318基因片段,克隆到PMD18-T载体上酶切鉴定正确.测序结果表明3株幽门螺杆菌(M0、 M13、11637)的HP0318序列大小均为756bp.序列分析显示在不同来源的HP菌株中表现出较高的保守性,最低相似性达到94%.结论 HP0318基因属于HP特异性基因,HP在沙鼠中适应性定植可能并不是由HP0318基因水平的变异引起,有必要进一步实验验证其功能和阐明其所具有的调控机制.

Cloning and characterization of HP0318 gene sequences of Helicobacter pylori after adaptive-colonization in Mongoliang gerbils

Objective To investigate the sequence characterization of HP0318 gene of Helicobacter pylori(HP) after adaptive-colonization in Mongoliang gerbils.Methods The gene encoding the HP0318 of HP was amplified from the chromosomal DNA of HP strain M0,NCTC11637and M13 by PCR.The M13 is a strain of adaptive-colonization in Mongolian gerbils abtained through multiple passage by M0 which is a clinical isolated strain.The 11637 is a type strain.The gene was cloned into pMD18-T-simple vector and then was sequenced.The gene sequence were referred to Genbank for BLASTn analysis and compared with DNASIS software.Results The HP0318 gene was successfully cloned into pMD18-T-simple vector.Sequence analysis indicated that the gene was very conservative in different HP strains.Its lowest nucleotide homology was 94%.Conclusion HP0318 is a peculiar gene of HP.The reason of adaptive colonization of HP may be not the genetic changes of HP0318.