基于HDA的超级细菌耐药基因NDM-1检测方法的建立

Establishment of detecting method of NDM-1 gene by the Helicase-dependent isothermal DNA amplification

本研究以建立解旋酶DNA恒温扩增(HDA)检测方法为目的,选择三种超级细菌NDM-1基因(metallo-beta-lactamase 1 gene)为目的基因,设计并合成特异性引物,建立和优化了可快速检测该种基因的HDA检测方法。特异性和灵敏度检测结果表明:该方法对NDM-1基因检测具有特异性,最低检测限可达10fg/μL,灵敏度高于普通PCR。提示HDA是一种针对NDM-1的快速、灵敏、便捷的非PCR核酸扩增技术。

In this paper,a new method,which based on Helicase-dependent isothermal DNA amplification (HDA) to detect NDM-1 (metallo-beta-lactamase 1 gene) of sample was established.A highly specific set of primers was synthesized to target the NDM-1 and amplificating temperature,amplificating time were optimized so as to establish Helicase-dependent isothermal DNA amplification method.Specific and sensitivity were tested which were compared with PCR method.The results shows that the sensitivity of HDA was 10 fg/μl of vector DNA which was higher than the result of PCR method.Detecting NDM-1 with HDA is specific and sensitive as well as PCR method and has lower instrumental requirement.