Genetic diversity analysis of grapevine rupestris stem pitting-associated virus from grapevine by colony PCR-SSCP and -RFLP

文献类型： 外文期刊

第一作者： Hu, Guo-Jun

作者： Hu, Guo-Jun;Zhu, Hong-Juan;Zhang, Zun-Ping;Fan, Xu-Dong;Ren, Fang;Dong, Ya-Feng

作者机构：

关键词： grapevine; grapevine rupestris stem pitting-associated virus (GRSPaV); RFLP; SSCP; genetic diversity analysis

期刊名称：ACTA VIROLOGICA （ 影响因子：1.827； 五年影响因子：1.632 ）

ISSN： 0001-723X

年卷期： 2022 年 66 卷 1 期

页码：

收录情况： SCI

摘要： We have developed methods for detecting the genetic diversity of grapevine rupestris stem pitting-associated virus (GRSPaV) based on restriction fragment length polymorphism (RFLP) and single stranded conformational polymorphism (SSCP) in the 905 nt 3' sequence. The amplicons were cloned from six grapevine cultivars, and colony polymerase chain reaction (colony PCR) using recombination bacteria was subsequently analyzed by RFLP and SSCP. Four haplotypes of SSCP and six haplotypes of Sac I RFLPs were defined. The two methods had a 40% discrepancy rate in showing the degree of diversity. All clones were sequenced and were used to construct a phylogenetic tree with seven previously reported GRSPaV sequences. In the tree, all the newly acquired sequences were divided into three clusters, I, II, and III, which corresponded to haplotypes I, II, and III of SSCP, respectively. Haplotype IV of SSCP was grouped into cluster II. A recombination analysis showed that haplotype IV has undergone a recombination event. Together, these results indicate that the SSCP assay is useful for the rapid identification of genetic diversity of GRS PaV. This is the first report of an analysis of the large fragment of GRSPaV by colony PCR-SSCP.

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