幽门螺杆菌适应性定植相关基因HP0318的克隆及序列分析

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

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。

Objective To investigate the sequence characterization of HP0318 gene of Helicobacter pylori（HP） after adaptivecolonization in Mongoliang gerbils. Methods The gene encoding the HP0318 of HP was amplified from the chromosomal DNA of HP strain M0, NCTCl1637and 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 13LASTn 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.