Dulieu P, Bar-Joseph M
S. Tolkowsky Laboratory, Department of Virology, Volcani Center, Bet Dagan, Israel.
J Virol Methods. 1989 Apr-May;24(1-2):77-83. doi: 10.1016/0166-0934(89)90009-8.
A simple method was developed to isolate dsRNA segments from disulphide crosslinked polyacrylamide gels. The dsRNA preparations from the P4 killer virus strain 77 of Ustilago maydis containing 7 genomic segments with molecular sizes ranging from 0.36 to 6.7 kbp, the 20 kbp dsRNA associated with the '447' cytoplasmic male sterility in Vicia faba and the 23 kbp genomic dsRNA of citrus tristeza virus (CTV) were separated on disulphide crosslinked polyacrylamide gels. After UV visualisation, the dsRNA bands were excised from the gel and dissolved using 2-mercaptoethanol. The dsRNA were then purified from the solubilized fractions by specific adsorption on microgranular cellulose and elution with a small volume of water. The method is rapid, simple and convenient for the isolation of all the tested dsRNAs segments.
开发了一种从二硫键交联的聚丙烯酰胺凝胶中分离双链RNA片段的简单方法。从玉米黑粉菌的P4杀手病毒株77制备的双链RNA制剂含有7个基因组片段，分子大小范围为0.36至6.7 kbp，与蚕豆“447”细胞质雄性不育相关的20 kbp双链RNA以及柑橘衰退病毒（CTV）的23 kbp基因组双链RNA在二硫键交联的聚丙烯酰胺凝胶上进行了分离。紫外线可视化后，从凝胶中切下双链RNA条带，并用2-巯基乙醇溶解。然后通过在微颗粒纤维素上特异性吸附并用水少量洗脱，从溶解的部分中纯化双链RNA。该方法对于分离所有测试的双链RNA片段而言快速、简单且方便。
