斑点叉尾鮰疱疹病毒PCR-ELISA检测方法的建立  

Development of a PCR-ELISA Assay for the Detection of the Channel Catfish Virus

作　　者：杜玉东[1] 陈孝煊[1] 吴志新[1] 王敏[1] 刘迁[1] 夏君[1] 王树云[1] 李莉娟[1] 

机构地区：[1]华中农业大学水产学院,武汉430070

出　　处：《华中农业大学学报》2009年第5期604-608,共5页Journal of Huazhong Agricultural University

基　　金：国家自然科学基金(30700624);"十一五"国家科技支撑计划(2006BAK02A22);农业部948项目(2005-Z37);华中农业大学科技创新基金资助

摘　　要：针对斑点叉尾鮰疱疹病毒(CCV)的ORF6基因设计引物和探针,PCR产物在扩增过程中标记地高辛,探针5′端用生物素标记,建立了CCV的敏感、快速、环保的PCR-ELISA检测方法。对反应条件进行了优化,并评估了该方法的特异性和敏感性。结果表明:该方法可特异性检测CCV DNA,与各对照均无交叉反应,具有高度特异性;该方法对CCV DNA的检测阈值为5 fg,敏感性是常规PCR检测方法的10倍。用此方法对人工感染样品进行检测,能够检测到感染组织中的CCV DNA。该方法能够定性和半定量检测CCV DNA,可用于斑点叉尾鮰疱疹病毒的检测、斑点叉尾鮰暴发性流行病的诊断。A rapid and sensitive PCR-ELISA assay was developed for detection of channel catfish virus （CCV）. The specific 135 bp fragment labeled with digoxigenin was amplified from the ORF6 gene of CCV. Approximate 5 fg of purified DNA could be detected by the PCR-ELISA system, indicating this method had high sensitivity,10 times as much as which detected by conventional PCR. No cross-reaction was observed against the controls. The result in this study showed that this sensitive and specific method was useful for early detection of CCV infection.

关 键 词：PCR-ELISA 斑点叉尾鮰疱疹病毒 敏感性 特异性 

分 类 号：Q939.4[生物学—微生物学]