关键词: 葡萄茎痘伴随病毒, RT-PCR, nad5, 检测

Abstract: This study prefer to establish a rapid, sensitive, accrual and practical detection system for Grapevine rupestris stem pitting associated virus (GRSPaV) based on internal control. These grapevine varieties such as‘Red Golbe’ that were detected to carry GRSPaV by RT-PCR were selected as testing material. Using SDS method to extract high quality total RNA from grape leaves and phloem, which was template to synthesize the cDNA by the guide of GRSPaV special antisense primer. After that, through PCR amplification, the 830 bp special fragmentwas gained. The co-amplification system of GRSPaV and plant mitochondrial nad5 as internal controlwas established. The use of internal control minimizes the risk of obtaining false negative RT-PCR results and avoids the need to eliminate contaminating DNA in extracts. The GRSPaV special fragment and nad5 aim fragmentwere cloned and sequenced respectively. The sequenced results aligned with nucleotide array in NCBI. The aligned results showed that GRSPaV has 96% similarity with array code
AF057136 and nad5 has 96.67% similarity with array code D37958.

Key words: Grapevine rupestris stem pitting associated virus (GRSPaV), RT-PCR, nad5, Detection