幽门螺杆菌基因分型及耐药检测寡核苷酸芯片的制备及鉴定

Preparation and Identification of the Oligonucleotide Chip for Detecting the Virulence-associated Genotypes and Drug Resistance of Helicobacter Pylori

目的探讨应用寡核苷酸芯片简单快速检测幽门螺杆菌(HP)感染并对其致病性相关的基因型及耐药性同时进行检测的方法,并进行鉴定。方法根据Hp的致病性将其分为cagA+和cagA-两种基因型,通过检测23SrRNA基因是否发生点突变判断其对克拉霉素的耐药性。应用寡核苷酸芯片技术对临床标本的Hp进行基因分型和耐药性检测,并制备相应的寡核苷酸芯片。通过构建的野生型模板和突变模板证实制备的寡核苷酸芯片的准确性。结果制备的寡核苷酸芯片可准确地将Hp分为cagA+和cagA-两种基因型,并能够通过检测23SrRNA基因中4种常见的、与克拉霉素耐药性密切相关的点突变,判断幽门螺杆菌对克拉霉素的耐药性。结论应用寡核苷酸芯片可快速检测Hp感染并同时对其致病性相关的基因型及耐药性进行检测。

Objective To develop an oligonucleotide microarray-based method for the simultaneous detection of Helicobacter pylori （Hp） infection, virulence-associated genotypes, and drug resistance. Methods Hp was classified into cagA ＋ and cagA- genotypes based on its virulence. Clarithromycin-resistance of Hp was identified by to simultaneously with clarithromyc existence of point mutations in 23S rRNA. We constructed an oligonucleotide microarray chip diagnose Hp infection and detect its virulence-associated genotypes and mutations associated in-resistance. The diagnostic accuracy of the constructed microarry was tested with templates of wild type and mutated type. Results The oligonucleotide chip accurately detected cagA ＋ and cagA- geno- types of Hp, as well as four common point mutations in 23S rRNA related to clarithromycin-resistance. Conclusion Oligonucleotide microarry chip can be used to diagnose Hp infection and test its virulence-associated genotypes and drug resistance simultaneously.