番茄黄化曲叶病毒的实时荧光定量PCR检测

Real-time PCR for Quantification of Tomato yellow leaf curl virus in Tomato Plants

番茄黄化曲叶病毒病(Tomato yellow leaf curl virus disease,TYLCVD)是目前为害番茄生产的重要病害,准确检测TYLCV含量是深入研究该病毒致病机理以及抗病育种的基础。根据TYLCV的DNA保守序列设计引物,基于SYBR Green I检测模式,构建了TYLCV实时荧光定量PCR检测体系,并且对接种病毒30d后的感病番茄植株中的TYLCV进行检测。结果显示,1ng·μL-1感病番茄总DNA中约含有3.47×106个病毒拷贝。利用实时荧光定量PCR法比普通PCR法的灵敏度提高100倍。

Tomato yellow leaf curl virus disease(TYLCVD)is one of the most devastating plant diseases in the world.Accurate quantification of TYLCV is the foundation for further studying the pathogenesis of this virus and also for resistance breeding.This paper constructed a real-time PCR testing system to quantify TYLCV in tomato by the primer,designed on the conservative sequences of TYLCV complete genome,and a testing mode based on SYBR Green I technology.The TYLCV were detected in the infected tomatoes after 30 days post-inoculation,about 3.47×10 6virus copies could be detected in 1 ng·μL-1 total DNA in the infected tomato plants.This method is 100-fold more sensitive than the traditional PCR.