重组酶聚合酶等温扩增快速检测结核分枝杆菌

Rapid detection of Mycobacterium tuberculosis using recombinase polymerase amplification

目的建立一种基于重组酶聚合酶介导的等温扩增技术(RPA)快速检测结核分枝杆菌的方法。方法根据结核分支杆菌基因保守序列设计的2对引物和特异性寡核苷酸探针,以结核分枝杆菌阳性菌株和其他5种非结核分枝杆菌基因组DNA为模板,考察该方法的检测灵敏度和特异性。结果 RPA快速检测结核分枝杆菌方法仅需15 min,检测灵敏度为可检出0.043 ng/μl的基因组DNA;5种非结核分支杆菌均不能扩增,特异性较高。结论本研究建立了结核分枝杆菌的RPA快速检测方法,具有快速、简单、成本较低等优点,为结核分枝杆菌的快速检测提供一个新的工具。

Objective To develop a rapid detection method of Mycobacterium tuberculosis using recombinase polymerasemediated isothermal amplification（RPA）. Methods Two pairs of primers and specific oligonucleotide probes were designed according to the conserved sequence of Mycobacterium tuberculosis. The analytical sensitivity and specificity of RPA method were also investigated. Results The detection time of RPA method for Mycobacterium tuberculosis was only15 min. The analytical sensitivity of this method was 0.043 ng / μl genomic DNA of Mycobacterium tuberculosis strain. No amplified was detected from 5 other microorganisms. Conclusion The RPA rapid detection method of Mycobacterium tuberculosis has the advantages of rapid, simple and low cost, which provides a new tool for the routine detection.