病毒重组细菌人工染色体构建、突变及应用

Viral bacterial artificial chromosomes: construction, mutagenesis and application

如何快速、方便地保存和修饰基因组较大的DNA和RNA病毒核酸一直是病毒学研究的重点。细菌人工染色体(bacterial artificial chromosome,BAC)为解决这两个问题提供了便利,它可以容纳300 kb的基因序列,能够通过大肠杆菌诱变技术进行有效修饰,目前已经成为编辑大基因组病毒基因的有效工具。DNA病毒,如疱疹病毒和痘病毒的基因组已被克隆到BAC载体上。此外,一些RNA病毒,如冠状病毒和黄病毒科病毒等也建立了基于BAC的反向遗传操作系统。现概述不同病毒重组BAC构建策略,总结了常用的对BACs进行基因修饰的技术,介绍了BAC载体重组病毒的拯救以及将BAC序列从病毒基因组中删除的方法,简述了病毒重组细菌人工染色体的应用。

Maintenance and manipulation of large DNA and RNA virus genomes have been a focus for virological research. Bacterial artificial chromosome（BAC） vectors provids a solution as they can harbor large DNA sequences and can efficiently be modified using well-established mutagenesis techniques in Escherichia coli, and has become an effective tool for editing large genomes. Numerous DNA virus genomes of herpes virus and pox virus were cloned into BAC vectors. In addition, several reverse genetic systems for RNA viruses such as members of Coronaviridae and Flaviviridae could be established based on BAC constructs. In this paper, we provide an overview on the strategies used for the generation of virus BAC vectors and also on systems currently available for various virus species. Furthermore, we address common mutagenesis techniques that allow modification of BACs and review the reconstitution of viruses from BAC vectors and strategies of removing the bacterial sequences from the virus genome. Finally, we briefly discuss the application of viral bacterial artificial chromosomes.