马铃薯S病毒的RT-PCR检测

Detection of Potato Virus S by Reverse Transcription-polymerase Chain Amplification

根据马铃薯S病毒(PVS)的外壳蛋白基因序列,设计合成了一对寡核苷酸引物。以感染PVS的马铃薯组织和健康的组织为材料,对提取植物总RNA的两种方法进行了比较,并对总RNA的提取方法进行改进,获得了纯度较高,完整性较好的总RNA。以此为模板,进行cDNA合成及PCR扩增,从感病组织扩增得到一段长度约642bp的特异PCR扩增产物,与理论设计的外壳蛋白基因大小一致,而健康组织无此扩增产物。从而建立了检测PVS快速灵敏简便的新方法,在基因水平上为PVS的检测提供了新手段。

A pair of DNA primers were designed and synthesized based on the nucleotide sequence of the coat protein （CP） gene of potato virus S （PVS）. Two methods were compared for isolation of total RNA from virus-infected and healthy potato tissue. The total RNA with high purity and quality was obtained using the improved method. The RNA could be used for cDNA synthesis and polymerase chain reaction （PCR）. A specific PCR fragments about 642 bp was obtained, which was same as the length of PVS CP gene, while no amplified products were obtained from the healthy tissue samples. The experiment established a rapid, sensitive and simple means for PVS detection and provided a new method for PVS detection at the level of gene.