甘薯潜隐病毒RT-PCR检测方法的建立及初步应用  

Establishment and Application of RT-PCR Assay for Detection of Sweet Potato Latent Virus

作　　者：张小梅[1] 侯文邦[1] 李广录[2] Zhang Xiaomei;Hou Wenbang;Li Guanglu(College of Agronomy,Henan University of Science and Technology,Luoyang,471003;Animal Science and Technology College,Henan University of Science and Technology,Luoyang,471003)

机构地区：[1]河南科技大学农学院,洛阳471003 [2]河南科技大学动物科技学院,洛阳471003

出　　处：《分子植物育种》2023年第2期544-550,共7页Molecular Plant Breeding

基　　金：河南省科技惠民专项(182207310015)资助。

摘　　要：甘薯是中国重要的旱地作物,具有天然的杂交优势,适应能力强且营养丰富,在中国广泛种植。在甘薯种植过程中,甘薯病毒的感染会导致甘薯产量与品质的下降,其中甘薯潜隐病毒(sweet potato latent virus,SPLV)是甘薯生产上常见的病毒之一,建立甘薯潜隐病毒的快速检测方法,对于该病的诊断及防御极其重要。根据NCBI数据库中甘薯潜隐病毒的全序列,选择保守性较强的外壳蛋白基因(cp基因),设计了一对全长引物与两对特异性引物。利用全长引物构建了甘薯潜隐病毒的阳性标准质粒,完成了特异性引物的筛选,并对其扩增条件进行了退火温度、循环数及引物浓度的优化。对该方法的灵敏度及特异性进行了检测,确定了甘薯潜隐病毒的RT-PCR检测技术体系。利用本体系对采集的15份疑似染病的甘薯样本进行检测,检测结果中有3份为甘薯潜隐病毒阳性。试验证明该检测方法具有很高的特异性与灵敏性,能够在甘薯脱毒薯苗生产中甘薯潜隐病毒的检测提供有力的技术支持。Sweet potato is an important dry land crop in China. It has natural heterosis, strong adaptability and rich nutrition, so it is widely planted in China. In the process of sweet potato planting, sweet potato virus infection will lead to the decline of sweet potato yield and quality. Sweet potato latent virus is one of the main virus which reduce in sweet potato viruses. To establishing a rapid detection method of SPLV is very important for the diagnosis and defense of the disease. According to the published complete sequences of SPLV cp gene, three pairs of primers was designed and synthetized. We constructed the positive standard plasmid of sweet potato latent virus with full-length primers and the selecting of special primers was completed. The detection procedure of reverse transcription-polymerase chain reaction(RT-PCR) for SPLV was built in this paper based on the optimization of the annealing temperature, number of cycles, primers density, sensitivity and specificity. Using the RT-PCR method clinical 15 samples, 3 samples were positive. The result indicates that the RT-PCR is very high specificity and sensibility, it can provide technical support for the rapid detection of SPLV virus in sweet potato.

关 键 词：甘薯 甘薯潜隐病毒 CP基因 RT-PCR 

分 类 号：S435.31[农业科学—农业昆虫与害虫防治]