江苏水稻黑条矮缩病病毒的RT-PCR分析和快速检测

Detecting and Analysis of Rice Black Stripe Dwarf Virus(RBSDV) on Rice in Jiangsu Province with RT-PCR

为了明确江苏新发生的矮缩病害为水稻黑条矮缩病,采用RT-PCR和序列测定方法对其病原进行了精确鉴定。根据水稻黑条矮缩病病毒基因组序列的信息,设计扩增RNA聚合酶基因、外壳蛋白基因和RNA沉默抑制子基因以及核心蛋白基因部分编码区的4对引物,提取感染水稻黑条矮缩病病毒的水稻植株总RNA并进行RT-PCR扩增,可以在感病植株中分别扩增出水稻黑条矮缩病病毒基因组特有的4个条带,测序结果表明,来源于江苏南京的分离物的4个基因部分编码片段与已登录的对应基因片段核苷酸序列同源性达99%以上,共发现7个碱基的突变。经生物信息学分析发现7个单碱基突变都是同义突变,氨基酸序列没有改变。根据以上结果确认该病病原为水稻黑条矮缩病病毒。利用该方法可以对水稻和其他寄主进行早期病毒检测。

To determine the pathogen of rice dwarf disease which has caused yield loss in Jiangsu Province in recent years was caused by rice black stripe dwarf virus（RBSDV）. Four pairs of primers to amplify RNA dependent RNA polymerase and the coat protein and core protein and silencing suppressor of RBSDV were designed. Four bands with about 200 bp to 300 bp were obtained in infected rice plants by RT-PCR, respectively. The separate segment was ligated to pGEM-T-easy vector,then sequenced with M13 primers. Their identities to rice black-streaked dwarf virus were higher than 99 % and seven synonymous substitution were found. The results showed that the virus which causes rice dwarf disease in Jiangsu is RBSDV. The identification of RBSDV in the small brown planthopper（ Laodelphax striatellus ）, rice and other host plants is of great importance for the disease forecasting. This approach can be used for early detection of RBS- DV in infected rice and other host plants.