唐菖蒲2种主要病毒的多重RT-PCR检测

摘要

根据黄瓜花叶病毒(cucumber mosaic virus,CMV)和烟草花叶病毒(tobacco mosaic virus,TMV)外壳蛋白基因序列分别设计特异性引物对,运用单重RT-PCR检测到病毒并筛选出适合多重RT-PCR的引物对.以唐菖蒲18S rRNA为内参照,建立同时检测CMV和TMV的多重RT-PCR方法.此方法可从带病样品中扩增出CMV( 629bp)和TMV(423 bp)2条特异片段.测序结果表明:CMV扩增产物与其他植物分离物核苷酸的同源性为93% ～97%,TMV扩增产物与其他分离物核苷酸的同源性可达99%.%Specific primer pairs were designed according to the gene sequence of cucumber mosaic virus( CMV ) and tobacco mosaic virus (TMV) coat protein region. The two viruses could be detected by simple RT-PCR with the primers and the suitable primers were selected for multiplex RT-PCR. Using 18S rRNA of gladiolus as an internal control, a multiplex RT-PCR program was established to simultaneously detect the two viruses of Gladiolus hybridus. Two specific bands,CMV(629 bp)and TMV(423 bp)were amplified from all the samples infected with CMV and TMV by the multiplex RT-PCR. Sequence analysis of the amplified products showed that the nucleotide sequences homology of CMV and TMV was 93% ~97% and 99% , respectively, compared with the sequences of other isolates.