陕北地区马铃薯卷叶病毒的RT―PCR检测与序列分析.docVIP

陕北地区马铃薯卷叶病毒的RT―PCR检测与序列分析 　　摘要：以陕北8个县（区）种植2～3代疑似带毒的马铃薯叶片为材料，用Trizol法提取马铃薯叶片总RNA，以已公布的马铃薯卷叶病毒外壳蛋白（CP）基因序列设计特异引物，通过RT-PCR扩增目的DNA片段，对CP基因进行克隆和测序，采用DNAstar软件分析序列的一致性。结果表明，从陕北8个县（区）马铃薯叶片中均扩增到与预期大小一致、长度为336 bp的目的片段，说明马铃薯均感染了卷叶病毒。陕北8个县（区）马铃薯卷叶病毒CP基因之间的序列一致性达99.6%以上，与国内外其他地区10个样品CP基因的序列一致性为96.2%～99.8%，表明马铃薯卷叶病毒的CP基因序列比较保守。 　　关键词：马铃薯卷叶病毒；CP基因；RT-PCR检测；序列分析 　　中图分类号：S435.32 文献标识码：A 文章编号：0439-8114（2014）19-4734-03 　　DOI：10.14088/j.cnki.issn0439-8114.2014.19.060 　　RT-PCR Detection and CP Gene Sequence Analysis of Potato Leaf Roll Virus in Northern Shaanxi Province 　　FENG Guang-hui，DU Hu-ping，LI Xia-long，KANG Fu-ren 　　（College of Life Science，Yulin University， Yulin 719000，Shaanxi，China） 　　Abstract： Taking 2-3 generations of suspected poisonous potato leaf planted in eight counties （districts） of Northern Shaanxi province as materials， the total RNA was extracted from potato leaves by Trizol method. Specific primers were published coat protein（CP） gene sequence of potato leaf roll virus. The DNA fragment was amplified by RT-PCR. Then the CP gene was cloned and sequenced. Sequence identity was analyzed by DNAstar software. The results showed that the gene sequence amplified from potato leaves from 8 counties in Northern Shaanxi area were consistent with the expected size with length of 336 bp. These potatoes were all infected with potato leaf roll virus. The CP gene sequence identity between potato leaf roll virus in 8 counties （districts） of Northern Shaanxi was more than 99.6%. 10 samples of CP gene sequences from other regions at home and abroad had identity ranged from 96.2% to 99.8%. It is indicated that the CP gene of potato leaf roll virus is more conservative. 　　Key words： potato leaf roll virus； CP gene； RT-PCR detection； sequence analysis 　　马铃薯卷叶病毒（Potato leaf roll virus，PLRV）是马铃薯主要的病毒之一，分布于世界各地马铃薯种植区。卷叶病毒可通过种薯、蚜虫或嫁接传播，感染卷叶病毒会造成马铃薯减产30%～90%[1]。近年来，脱毒马铃薯在全国各地大面积推广，但随着脱毒种薯种植代（年）数的增加，病毒在植株体内不断积累和传播，导致马铃薯产量和品质逐渐下降，而且还会感染其他种类的病毒和类病毒。因此，及时、准确地检测田间种植马铃薯的带毒情况，采用茎尖分生