Pugachev Konstantin V, Guirakhoo Farshad, Ocran Simeon W, Mitchell Fred, Parsons Megan, Penal Caroline, Girakhoo Soheila, Pougatcheva Svetlana O, Arroyo Juan, Trent Dennis W, Monath Thomas P
Acambis, Inc., Cambridge, Massachusetts 02139, USA.
J Virol. 2004 Jan;78(2):1032-8. doi: 10.1128/jvi.78.2.1032-1038.2004.
Three consecutive plaque purifications of four chimeric yellow fever virus-dengue virus (ChimeriVax-DEN) vaccine candidates against dengue virus types 1 to 4 were performed. The genome of each candidate was sequenced by the consensus approach after plaque purification and additional passages in cell culture. Our data suggest that the nucleotide sequence error rate for SP6 RNA polymerase used in the in vitro transcription step to initiate virus replication was as high as 1.34 x 10(-4) per copied nucleotide and that the error rate of the yellow fever virus RNA polymerase employed by the chimeras for genome replication in infected cells was as low as 1.9 x 10(-7) to 2.3 x 10(-7). Clustering of beneficial mutations that accumulated after multiple virus passages suggests that the N-terminal part of the prM protein, a specific site in the middle of the E protein, and the NS4B protein may be essential for nucleocapsid-envelope interaction during flavivirus assembly.
对四种针对1至4型登革病毒的嵌合黄热病病毒 - 登革病毒（ChimeriVax - DEN）候选疫苗进行了三轮连续的蚀斑纯化。在蚀斑纯化以及在细胞培养中进行额外传代后，通过一致性方法对每个候选疫苗的基因组进行测序。我们的数据表明，用于启动病毒复制的体外转录步骤中使用的SP6 RNA聚合酶的核苷酸序列错误率高达每复制一个核苷酸1.34×10⁻⁴，而嵌合体用于在受感染细胞中进行基因组复制的黄热病病毒RNA聚合酶的错误率低至1.9×10⁻⁷至2.3×10⁻⁷。多次病毒传代后积累的有益突变的聚类表明，prM蛋白的N端部分、E蛋白中间的一个特定位点以及NS4B蛋白可能对于黄病毒组装过程中的核衣壳 - 包膜相互作用至关重要。
