缺口连接酶链反应检测沙眼衣原体DNA的实验研究

Detection of Chlamydia trachomatis DNA by gap ligase chain reaction (G-LCR)

目的 :建立缺口连接酶链反应方法检测沙眼衣原体。方法 :根据编码CT主要外膜蛋白的基因 (ompl)的稳定区设计2对互补的寡核苷酸探针。采用G -LCR和常规PCR扩增CT标准株DNA ,并对二者检测CT的敏感性进行比较。结果 :对5种CT标准株进行G -LCR扩增 ,均出现 5 4bp长度的DNA片段。敏感性实验可检测出 2个CT原体 ,而PCR可检测出2 0EBs。G -LCR较PCR敏感 10倍 ;在特异性方面 ,不扩增肺炎衣愿体及其他细菌DNA ,具有很高的特异性。结论 :G -LCR用于检测沙眼衣原体特异、敏感、快速、准确 ,可为CT感染的临床诊断提供依据。

Objective:To develop a new nucleic amplication method for detection of Chlamydia trachomatis DNA by gap ligase chain reaction(G-LCR).Methods:A G-LCR DNA amplification assay that targeted the outer major membrane protein gene(omp1)of CT was established to detect CT infection.The sensitivity and specificity of a newly developed G-LCR test was examined by the use of highly purified elementary bodies (EBs).DNA fragments of different species and from other bacteria1 were detected with G-LCR and routine polymerase chain reaction (PCR).Results:Using G-LCR,DNA fragments of 54bp were amplified from five different species.The sensitivity could be improved to detect out 2 chlamydial elementary bodies.G-LCR detected ten-fold EBs than PCR.No signal was observed when C.pneumoniae and other bacteria were used as templates.Conclusion:G-LCR is sensitive,rapid and specific for detection of Chlamydia trachomatis.