用聚合酶链反应-银染单链构象多态性鉴别腮腺炎病毒株

Differentiation of mumps virus strains by ploymerase chain reaction and silver stained singlestrand conformation polymorphism analysis

用腮腺炎病毒小疏水蛋白（ＳＨ）基因及其旁侧区的逆转录（ＲＴ）套式聚合酶链反应（Ｎ－ＰＣＲ）产物进行银染单链构象多态性（ＳＳＣＰ）分析，建立了较适的样品变性和聚丙烯酰胺凝胶电泳（ＰＡＧＥ）条件。根据单链电泳模式可将所测６株腮腺炎病毒分为四种ＳＳＣＰ模式，３株兰州野毒株Ｗｌｚ１，Ｗｌｚ２，Ｗｌｚ３呈现一种模式，这３株病毒的ＳＨ基因及其旁侧区ｃＤＮＡ序列之间平均差异仅为０．９６％；两株上海野毒株Ｗｓｈ１和Ｗｓｈ２株的相应序列之间差异为４％，呈现另两种模式。Ｅｎｄｅｒｓ株呈现一种与野毒株差别较大的特殊模式。３株兰州野毒与２株上海野毒相应区域序列之间差异为３．４％～４．５％，故本法可区别ＳＨ基因及其旁侧区ｃＤＮＡ序列之间差异为３．４％的野毒株，且ＳＳＣＰ模式的相近程度与相应区域中核苷酸序列同源性大小一致。

The cDNA fragments amplified through nested RT-PCR from the small hydrophobic (SH) protein gene and its flanking region of the five wild mumps viruses and Enders strain were subjected to singlestrand conformation polymorphism (SSCP) analysis by using silver staining. These five wild isolates could be divided into three SSCP patterns and they were differentiated from the Enders strain. Two wild strains which have 3.4% heterology of the nucleotide acid sequence could be distinguished by the method. The results were well correlated with sequence analysis of the SH gene segments, indicating high applicability of the silver staining SSCP analysis for differentiation of mumps virus strains. Furthermore, the method, which is not only quick, sensitive, safe and reliable, but also reproducible and inexpensive, can be used in the molecular epidemiology study of mumps viruses.