马铃薯纺锤块茎类病毒RT-PCR检测及全序列分析

Detection of Potato Spindle Tuber Viroid by RT-PCR and Analysis of Its Complete Sequences

根据马铃薯纺锤块茎类病毒(PSTVd)基因序列设计合成引物,以感病组织和健康组织总RNA为模板,经RT PCR法扩增出全长的cDNA片段,结果从感病组织中扩增出与预期的360bp大小的目标片段,而健康组织无此扩增产物;将其克隆到质粒pGEM T载体上,进行全序列分析。结果表明,与国内外的报道相比较,核苷酸同源率高达98%以上。PCR产物克隆作为RT PCR反应的阳性对照,解决了毒源保存和传播的问题,为进一步进行抗PSTVd基因工程研究打下了良好基础。

A pair of primers were designed and synthesized based on PSTVd gene. The excepted size 360bp was amplified by RT-PCR from the infected samples, while no amplified products were obtained from the healthy tissue samples. The unique amplified product was then cloned into the pGEM-T vector and sequenced. Comparison of the civil and foreign nucleotide sequence showed that the homology are 98%. Acting as the positive control of RT-PCR, the clone of PCR product helped resolve the problem of storing the viroid source and preventing it from spreading. This paved a path for breeding the transgenic potato resistant to PSTVd.