柑橘衰退病、裂皮病和碎叶病的多重RT-PCR检测方法研究  

Simultaneous detection of Citrus tristeza virus(CTV),Citrus exocortis viroid(CEVd)and Citrus tatter leaf virus(CTLV) by multiplex RT-PCR

作　　者：肖远辉[1] 曾继吾[2] 张秋明[1] 周碧容[2] 

机构地区：[1]湖南农业大学园艺园林学院,长沙410128 [2]广东省农业科学院果树研究所,广州510640

出　　处：《植物病理学报》2007年第1期31-35,共5页Acta Phytopathologica Sinica

基　　金：广东省科技重点计划资助项目(2003B21601)

摘　　要：本研究以广东省农业科学院果树研究所隔离网室内通过嫁接分别感染CTV、CEVd和CTLV的柑橘树皮为实验材料,建立了以oligo(dT)为反转录引物的RT-PCR检测体系,成功检测到在病毒RNA 3′末端带有ploy_(A)加尾的CTV和CTLV。实验过程中,发现不具有ploy_(A)加尾的环状类病毒CEVd,同样可以用oligo(dT)反转录的RT-PCR检测出来,通过测序分析,其同源性在96%以上,并发现在CEVd序列中有一个富含A的区段。在此基础上,进一步研究了同时检测CTV、CEVd和CTLV的多重RT-PCR检测体系,该方法能为这3种病害的检测简化步骤、节省时间、降低成本。By use of an oligo（dT） 18 as a reverse transcription primer, polyadenylated RNA viruses CTV and CTLV were detected by RT-PCR. CEVd could be also detected in this way. CEVd is a non-polyadenylated RNA viroid, but there is a fragment rich in A in CEVd sequence. On the bases of uniplex RT-PCR, a multiplex RT-PCR was developed to detect CTV, CEVd and CTLV simultaneously. The results of the multiplex RT-PCR to detect each of the pathogens were consistent with uniplex RT-PCR. The multiplex RT-PCR provides a simple and rapid method for detecting CTV, CEVd and CTLV in citrus plants in a large number at the same time.

关 键 词：柑橘衰退病毒(CTV) 柑橘裂皮病类病毒(CEVd) 柑橘碎叶病毒(CTLV) 多重RT—PCR oligo(dT)18 

分 类 号：S436.661.1[农业科学—农业昆虫与害虫防治]