关键词: 葡萄, 葡萄卷叶伴随3型病毒, 葡萄A病毒, 病毒外壳蛋白基因, 病毒RNA沉默抑制子基因, RT-PCR多重检测, 病毒变异株

Abstract: Co-infection of Grapevine leafroll associated virus-3（GLRaV-3）and Grapevine virus A（GVA）was a common phenomenon on some cultivars of Vitis vinifera L. × Vitis labrusca L. in Sichuan Province. Semi-quantitative RT-PCR and qRT-PCR were used to investigate the relative accumulations of the aforementioned two viruses in different tissues of infected grape trees（Vitis vinifera L. × V. labrusca L. ‘Himrod’）. The results suggested the relative accumulations of viruses in petiole and phloem were higher than that of others tissues. The multiple RT-PCR was developed to amplify the target genes of GLRaV-3 and GVA in phloem tissues of mature shoots（Vitis vinifera L. × V. labrusca L.‘Fujiminori’）simultaneously. The full length of coat protein gene（CP）of GLRaV-3 and partial genome region of GVA（including partial sequence of CP and full length of viral suppressor of RNA silencing gene，VSR）were cloned from V. vinifera L. × V. labrusca L.‘Fujiminori’separately. The nucleotide homology analysis and phylogenetic studies on different isolates of each virus were possibly revealed that CP from different GLRaV-3 isolates was highly conserved，whereas GVA Sichuan isolate contained very divergent variants and thus it could be quasispecies.

Key words: grape, Grapevine leafroll associated virus-3, Grapevine virus A, coat protein, virus silencing suppressor, multiple RT-PCR, variant