对虾白斑综合症病毒变性高效液相色谱检测方法的建立与应用  

Establishment and Application of Denaturing High-performance Liquid Chromatography for White Spot Syndrome Virus

作　　者：赵巍[1,2] 马亚[1,2] 赵玉然[2] 杨大伟[1,3] 梁成珠[2] 刘荭[4] 何俊强[4] 史秀杰 岳志芹[2] 

机构地区：[1]中国海洋大学,山东青岛266003 [2]山东出入境检验检疫局技术中心,山东青岛266002 [3]安徽农业大学,安徽合肥230036 [4]深圳出入境检验检疫局水生动物疫病国家重点实验室,广东深圳518010

出　　处：《中国动物检疫》2011年第5期47-50,共4页China Animal Health Inspection

基　　金：国家质量监督检验检疫总局科研项目(2007IK267)

摘　　要：据对虾白斑综合症病毒(WSSV)的基因保守序列,使用Primer Explorer V3软件设计了2条引物,利用PCR的扩增产物,结合变性高效液相色谱(DHPLC)技术,建立了一种对虾白斑综合症病毒的DHPLC快速检测方法。该检测方法特异性好,与传染性皮下和造血器官坏死病毒、斑节对虾杆状病毒、肝胰腺细小病毒、对虾杆状病毒以及对虾基因组DNA无交叉反应;检测灵敏度较高,约为10-5ng/μL,高于常规PCR及荧光定量PCR的灵敏度。用建立的DHPLC方法对100尾对虾样品进行了临床检测,结果与荧光定量PCR检测结果一致。WSSV的DHPLC检测方法,特异性强、灵敏度高,是对WSSV进行有效检测的一种新方法。Two specific primers were designed based on conservative gene of white spot syndrome virus （WSSV） using Primer Explorer V3 software. A detection method for WSSV using the denaturing high-performance liquid chromatography （DHPLC） was established. The WSSV LAMP assay did not react with infetious hypodermal and haematopoietic necrosis virus （IHHNV）, monodon baculovirus （MBV）, hepatopancreatic parvovirus （HPV）, baculovirus penaei （BP） or genome DNA of shrimp, indicating the good specificity. The detection limit of DHPLC assay was approximately 10.5 ng/μL WSSV DNA, which was higher than that of conventional PCR and the real-time PCR. 100 clinical shrimp samples were detected with the DHPLC method and the results were consistent with the real-time PCR method. The DHPLC assay was accurate, specific and sensitive, having great potential in diagnosis of WSSV infection.

关 键 词：变性高效液相色谱 对虾 白斑综合症病毒 检测 

分 类 号：S945.19[农业科学—水产养殖] S945.4[农业科学—水产科学]