5种病毒性脑炎相关RNA病毒多重RT-PCR检测方法的初步建立  

Preliminary development of multiplex RT-PCR method for five kinds of encephalitis-associated RNA viruses

作　　者：孔祥军[1] 申镇维[1] 吴蕊[1] 李宝辉[1] 陈虎[1] 何继红[1] 赵连利[1] 于庆杰[1] 姬春雪[1] 

机构地区：[1]河北省沧州市中心医院中心实验室,河北沧州061001

出　　处：《中国生物制品学杂志》2014年第1期115-119,共5页Chinese Journal of Biologicals

基　　金：河北省沧州市科学技术研究与发展计划(1123010ZD)

摘　　要：目的建立肠道病毒、乙型脑炎病毒、麻疹病毒、风疹病毒和腮腺炎病毒的多重RT-PCR检测方法。方法根据GenBank中登录的肠道病毒、乙型脑炎病毒、麻疹病毒、风疹病毒和腮腺炎病毒基因组序列,分别设计肠道病毒5′UTR基因、乙型脑炎病毒E基因、麻疹病毒M基因、风疹病毒E基因和腮腺炎病毒M基因的特异性引物,建立多重RT-PCR检测方法,并进行敏感性验证。结果应用5种引物可分别扩增脊髓灰质炎病毒、乙型脑炎病毒、麻疹病毒、风疹病毒和腮腺炎病毒的152、429、111、352和274 bp特异基因条带。对脊髓灰质炎病毒、乙型脑炎病毒、麻疹病毒、风疹病毒和腮腺炎病毒的的敏感度分别为62.5 CCID50/ml、250 PFU/ml、125 CCID50/ml、125 CCID50/ml和125 CCID50/ml。结论初步建立了5种脑炎RNA病毒多重RT-PCR检测方法,该法敏感性良好,可用于5种脑炎相关RNA病毒的快速检测。Objective To develop a muhiplex RT-PCR method for enterovirus, Japanese encephalitis （JE） virus, measles virus, rubella virus and mumps virus. Methods Specific primers for 5＇ UTR gene of enterovirus, E gene of JE virus, M gene of measles virus, E gene of rubella virus and M gene of mumps virus were designed according to the genome sequences of enterovirus, JE virus, measles virus, rubella virus and mumps virus reported in GenBank, based on which a multiplex RT-PCR method was developed and verified for sensitivity. Results Specific gene bands of poliomyelitis virus, JE virus, measles virus, rubella virus and mumps virus, at lengths of 152, 429, 111, 352 and 274 bp respectively, were amplified by using the five primers. The sensitivities of developed multiplex RT-PCR method for the above five kinds of viruses were 62. 5 CCID50/ml, 250 PFU/ml, 125 CCID50/ml, 125 CCID50/ml and 125 CCID50/ml respectively. Conclusion A multiplex RT-PCR method for five kinds of＇ encephalitis-associated RNA viruses was developed preliminarily, which showed high sensitivity, and might be used for the rapid detection of the viruses.

关 键 词：肠道病毒 乙型脑炎病毒 麻疹病毒 风疹病毒 腮腺炎病毒 病毒性脑炎 逆转录聚合酶链反应 

分 类 号：R373.31[医药卫生—病原生物学] Q789[医药卫生—基础医学]