以dsRNA为模板的梨脉黄病毒RT-PCR检测技术研究

Study on the RT-PCR Detection System for Pear vein yellow virus by Using dsRNA as Template

用已知带梨脉病毒(Pearveinyellowvirus,PVYV)的库尔勒香梨植株和库尔勒香梨无病毒实生苗为供试材料,分别选用新鲜幼嫩叶片、4℃条件下保存2～4d的幼嫩叶片、冷冻的幼嫩叶片和皮层为试材,进行dsRNA的分离提取实验,得到了与PVYV基因组RNA长度9306bp相当大小的dsRNA谱带,并以dsRNA为模板,利用设计的特异引物,研究建立了梨脉黄病毒(PVYV)的RT-PCR检测技术体系,能有效地扩增出PVYV的一条长为316bp的特异片段,该技术可广泛用于果树病毒的分子检测。

The double-strand RNA(dsRNA)from fresh,refrigerated(4℃2～4days),freezen leaf and bark of Korlaxiangli pear variety infected with Pear vein yellow virus(PVYV)and virus-free seedling were extracted respectively.PVYV dsRNA about 9306bp(PVYV genome length)was obtained.Using dsRNA as template,the RT-PCR detection system of PVYV based on dsRNA template with differential primers were developed.A316bp specific nucleotide fragment of PVYV was ef-fectively amplified by this system.The technology can be used for the molecular detection of fruit tree viruses comprehensive-ly.