基因芯片技术检测恙虫病东方体DNA的方法建立

Establishment of diagnostic assay for scrub typhus with microarray techniques

目的 研制用于东方体检测的基因芯片 ,建立快速、灵敏、特异的恙虫病诊断方法。方法 根据东方体 5 6kD外膜蛋白基因序列 ,设计群特异性引物及探针 ,制备寡核苷酸芯片。用Cy3标记的引物 ,利用不对称PCR技术扩增DNA片段 ,将扩增的DNA片段与芯片上的探针杂交 ,用荧光扫描仪检测并分析信号。结果 建立的基因芯片能够检测恙虫病东方体标准株DNA的特异性荧光信号。具有特异、灵敏、快速的优点。结论 成功制备的基因芯片有望应用于恙虫病东方体多种样本的检测 ,为恙虫病提供最为理想的诊断方法。

Aim To establish a rapid,sensitive and specific diagnostic assay for scrub typhus with microarray techniques.Methods Firstly,the specific primers and probe were designed according to the conservative and specific DNA sequence of 56kD protein of orientia tsutsugamush.Scecondly,the probes were spotted on glass slides to form microarrays.Then,the Cy3-labled Ot DNA fragment prepared by dissymmetical PCR were hybridized to the microarrays.Finally,the microarrays were scanned and analyzed with a scanner.Results The DNA microaray could detect the DNA of Ot standard strains of scrub typhus with adequate specificity and sensitivity.Conclusion The successfully developed DNA microarray might be widely applied in specific detection of various Ot samples and be a very useful method for diagnosis of Scrub typhus.