人型支原体实时荧光聚合酶链反应检测方法的初步建立

Establishment of real-time PCR assay to detect Mycoplasma hominis

摘要: 目的建立一种快速、灵敏和特异的人型支原体实时荧光聚合酶链反应(real-time PCR)检测技术。方法 依据人型支原体gap基因保守区域使用Beacon Designer 7.0软件设计引物和探针,建立人型支原体real-time PCR检测方法并优化。对优化后的方法进行实验室灵敏度、特异度和检测限评价,并与聚合酶链反应(PCR)进行比较。结果该real-time PCR对于人型支原体的检测限为50 cfu,PCR检测限为5103cfu,其检测灵敏度为PCR的100倍。所建立的检测方法对其他10种常见支原体、12种泌尿生殖道感染病原菌染色体及人类染色体的扩增均为阴性。结论本研究建立的real-time PCR方法可灵敏、特异的检测人型支原体,有望用于临床标本检测。

Abstract: Objective To develop a real-time PCR assay to detect Mycoplasma hominis. Methods By analyzing the gap gene of M. hominis, an optimized real-time PCR assay was designed. The specificity, sensitivity and detection limit of the assay were evaluated and compared with conventional PCR assay by using standard concentration DNA of M. hominis. Results The detection limit of the assay was about 50 cfu and the specificity of the assay appeared to be 100%. The sensitivity of this real-time PCR assay was 100 times higher than that of conventional PCR. Conclusion This real-time PCR assay might be a suitable method for the clinical detection of M. hominis.