半巢式RT-Realtime PCR检测南芥菜花叶病毒

Dectection of Arabis Mosaic Virus by Semi-Nested RT-Realtime PCR

根据南芥菜花叶病毒(Ar MV)外壳蛋白(CP)基因的保守序列,设计合成了3条巢式PCR引物和1条Taq MAN荧光探针,建立了半巢式RT-Realtime PCR检测Ar MV的新方法。该方法有机地结合了巢式PCR和Taq MAN探针检测技术。第二步半巢式-RT-Realtime PCR既是对第一步信号的进一步放大,也是对第一步PCR产物的确认,因此,检测的准确性、灵敏度比巢式PCR、Realtime PCR等方法高。结果表明:该方法检测灵敏度可达0.06 fg/μL植物总RNA。

Three nested primers and one TaqMAN probe were designed according to the CP gene of the Arabis mosaic virus ,with which we set up a new method of semi-nested RT-Realtime PCR to detect the ArMV.The nest-PCR and the probe-detection technique were combined maneuverably in this study.The semi-nested RT-Realtime PCR in the second step could both magnify the sigal and validate the result of the first step,which could provide a more sensitive and specific detection of the ArMV.The sensitivity of the method could reach 0.06 fg/μL of total plant RNA.