Lin B, Cotton R G, Trent D W, Wright P J
Department of Microbiology, Monash University, Clayton, Victoria, Australia.
J Virol Methods. 1992 Nov;40(2):205-18. doi: 10.1016/0166-0934(92)90069-p.
Genetic variation in 12 strains of dengue virus type 2, isolated from several epidemic areas in different years, was studied by chemical cleavage at mismatched cytosine in DNA:RNA heteroduplexes. End-labelled cDNA probes derived from the E and NS2A genes of the New Guinea C strain were hybridized to total RNA extracted from cells infected by individual isolates. Following modification of mismatched cytosine by hydroxylamine and nucleic acid strand cleavage by piperidine, the resulting fragments of radiolabelled probe were analysed by electrophoresis and autoradiography. The patterns of bands generated corresponded to the geographical groupings of the isolates. Thus this method is suitable in epidemiological studies for rapidly surveying a large number of isolates for genetic variation in a particular gene of interest.
通过对DNA:RNA异源双链体中错配胞嘧啶进行化学切割，研究了从不同年份的几个流行地区分离出的12株2型登革病毒的基因变异情况。将源自新几内亚C株E基因和NS2A基因的末端标记cDNA探针与从单个分离株感染的细胞中提取的总RNA进行杂交。在用羟胺修饰错配胞嘧啶并通过哌啶切割核酸链后，通过电泳和放射自显影分析放射性标记探针产生的片段。产生的条带模式与分离株的地理分组相对应。因此，该方法适用于流行病学研究，可快速检测大量分离株中特定感兴趣基因的基因变异。
