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A sensitive SYBR Green rt-qpcr method for grapevine virus E and its application for virus detection in different grapevine sample types

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Journal of Integrative Agriculture 2020年第7期19卷 1834-1841页

作者： REN Fang ZHANG Zun-ping FAN Xu-dong HU Guo-jun ZHANG Meng-yan DONG Ya-feng National Center for Eliminating Viruses from Deciduous Fruit Trees Research Institute of PomologyChinese Academy of Agricultural SciencesXingcheng 125100P.R.China

To develop a rapid and high-sensitivity method for detection of grapevine virus E(GVE),a SYBR Green based real-time fluorescence quantitative rt-pcr method(rt-qpcr)was *** method could be used to detect GVE specifically,and the sensitivity was about 100 times greater than conventional *** excellent linear correlation(R=0.997)and a high amplification efficiency(E=97.5%)were obtained from the standard curve of this *** tests revealed that the coefficients of variation in the intra-and inter-assay results were 0.31-1.03%and 0.82--262%,respectively,indicating a good *** rt-qpcr method could be used to detect GVE in a wide range of grapevine sample *** detection rates of rt-qpcr for nearly all sample types from different positions and seasons were higher than conventional *** detection rates in spring,summer,autumn and winter increased *** in autumn and winter were best for detection,and the detection rates of most samples were 80-100%,which were 10 to 40%higher than conventional *** general,old petioles and branches were the best tissues for GVE *** detection rates of these samples in each season were all 100%,which were 20 to 40%higher than conventional *** second highest rates were in the old leaf,with detection rates for rt-qpcr of 80-100%in all seasons,which were 20 to 40%higher than conventional *** could be difficultly detected in young leaves by conventional rt-pcr,and the detection rates were only 0-50%,while by rt-qpcr the rates could increase to 0--80%.A total of 33 out of 363 samples(belonging to 68 cultivars)from 20 regions in China were detected to be positive by rt-qpcr(9.1%),which was more than twice the rate of the conventional rt-pcr(3.9%).

关键词： grapevine grapevine virus E detection rt-qpcr conventional rt-pcr

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