运用限制性片段长度多态性分析方法鉴别麻疹疫苗株和野毒株病毒

Identification of Measles Vaccine Viruses and Wild Viruses by Restriction Fragment Length Polymorphism Method

目的应用和验证快速鉴别麻疹疫苗株与野毒株病毒的方法。方法采用逆转录-聚合酶链反应-限制性片段长度多态性分析(Reverse Transcription-Polymerase Chain Reaction-Restriction Fragment Length Polymorphism,RT-PCR-RFLP)方法,对四川省2009～2011年分离的麻疹疫苗株和野毒株病毒进行鉴定,同时与序列分析进行对比验证。结果 5株RT-PCR扩增阳性产物,经AflⅡ酶切后,1株麻疹疫苗株病毒被切成两个片段,分别为287碱基对(Base Pair,bp)和151bp;4株麻疹野毒株病毒均不能被AflⅡ酶切,仍呈单一条带。结论 RT-PCR-RFLP是一种快速、简便的鉴别麻疹疫苗株与野毒株病毒的方法。

Objective To apply restriction fragment length polymorphism （RFLP）method to quickly identify measles vaccine viruses and wild viruses. Methods Virus isolation were carried out by cell culture from urine and throat swab specimens collected from suspected measles cases during 2009- 2011 in Sichttan. Ribonucleic Acid （RNA） were extracted from the culture isolates. 438-base pair （bp） fragments targeting hemagglutinin gene of measles virus were amplified by reverse transcription- polymerase chain reaction （RT-PCR）, the resulting PCR products were then digested with restriction enzyme Aft II and visualized by capillary electropheresis. Results Of 5 positive RT-PCR products, one that proved to be measles vaccine virus by direct sequencing was digested by Aft II into 287-bp fragment and 151-bp fragment. The other four that proved to be measles wild viruses by direct sequencing could not be cut by Aft II and remained 438-bp fragments. Conclusion RT-PCR-RFLP is a rapid and simple method for identification of measles vaccine viruses and wild viruses.