Carman W F, Williamson C, Cunliffe B A, Kidd A H
National Institute for Virology, University of the Witwatersrand, Johannesburg, South Africa.
J Virol Methods. 1989 Jul;25(1):21-9. doi: 10.1016/0166-0934(89)90097-9.
A method is described whereby rubella virus RNA was reverse transcribed and the resulting cDNA enzymatically amplified using Taq polymerase. The reactions were carried out in a single reaction vessel, with only minor modifications to the buffer conditions between the reverse transcription and the subsequent amplification step. Using an oligonucleotide probe to the E1 glycoprotein region and limited restriction endonuclease mapping, the resulting amplified products were shown to be specific for rubella virus. This method was also successfully applied to crude cell lysates, without the need for RNA purification. The possible applications of the polymerase chain reaction as applied to RNA sequences are discussed.
本文描述了一种方法,通过该方法将风疹病毒RNA进行逆转录,并使用Taq聚合酶对所得的cDNA进行酶促扩增。反应在单个反应容器中进行,在逆转录和随后的扩增步骤之间,缓冲液条件仅需进行微小修改。使用针对E1糖蛋白区域的寡核苷酸探针和有限的限制性内切酶图谱分析,结果表明所得的扩增产物对风疹病毒具有特异性。该方法也成功应用于粗细胞裂解物,无需进行RNA纯化。文中还讨论了聚合酶链反应应用于RNA序列的可能用途。