钩端螺旋体赖型017鞭毛钩相关蛋白K基因的克隆及序列分析  

Amplification and Analysis of Flagella Hook-Associated Protein Gene of Leptospira Interrogans Seroval Lai

作　　者：于向华[1] 鲍朗[1] 谢勇恩 张会东 

机构地区：[1]四川大学华西医学中心基础与法医学院 感染免疫研究室,中国四川成都610041

出　　处：《生命科学研究》2002年第S1期101-107,共7页Life Science Research

基　　金：国家自然科学基金资助项目(30070670)

摘　　要：在以抑制消减杂交比较强毒株赖型钩端螺旋体017株和无毒株双曲钩体Patoc　I株　的基因组差异时,获得了一系列仅存在于强毒株而无毒株缺如的差异片段.选取差异片段AF325810设计特异性引物,以赖型钩端螺旋体017株基因组DNA为模板,进行巢式PCR扩增,PCR纯化产物T载体克隆,选取阳性克隆测序,进一步进行生物信息学分析,以获得强毒株赖型钩端螺旋体017株特有的毒力相关基因,DOT BLOT显示其在钩端螺旋体各株间有不同分布PCR扩增得到了产物为2kb大小的DNA片段,序列分析结果显示得到了问号钩端螺旋体赖型017株的鞭毛钩相关蛋白K基因的上游序列,为进一步探索钩端螺旋体的致病机制奠定了基础.A series of differential fragments which were found only in pathogenic Leptospira seroval lai when we compared the genomic difference of pathogenic Leptospira seroval lai and non-pathogenic Leptospira serova patoc I using subtractive suppressive hybridization(SSH) technique. According to the selected AF325810, the unknown region of this gene fragment is amplified by using nested PCR. The resulted product was purified and cloned into T vector,then the selected clone was sequenced and bioinformatics analyzed.Dot Blot shows there is different distributions among leptospira. A 2 kb DNA fragment was amplified. The sequence analysis showed that the upstream sequence of flagella hook-associated protein K gene was got, which laid solid foundation for further research in pathogenic mechanism of Leptospira.

关 键 词：钩端螺旋体 巢式PCR 鞭毛钩相关蛋白K(flgK) 生物信息学 

分 类 号：R346[医药卫生—基础医学]