Hashimoto Yoshitami, Costanzo Vincenzo
Clare Hall Laboratories, London Research Institute, EN6 3LD, Hertsfordshire, UK.
Methods Mol Biol. 2011;745:437-45. doi: 10.1007/978-1-61779-129-1_25.
A crucial process to ensure cell survival and genome stability is the correct replication of the genome. DNA replication relies on complex machinery whose mechanisms are being elucidated using different model systems. A major aspect of this process, which is an intense subject of investigation, is what happens when replication forks encounter obstacles impairing their progression such as modified bases, pausing sites, and single strand breaks. The detailed biochemical analysis of DNA replication in the presence of DNA damage has been impeded by the lack of a cell-free system recapitulating DNA replication. Here we describe assays based on the vertebrate Xenopus laevis egg extract to study the biochemical aspects of replication fork stability.
确保细胞存活和基因组稳定性的一个关键过程是基因组的正确复制。DNA复制依赖于复杂的机制，其原理正通过不同的模型系统进行阐释。这个过程的一个主要方面（也是一个深入研究的课题）是当复制叉遇到阻碍其前进的障碍时会发生什么，例如修饰碱基、暂停位点和单链断裂。由于缺乏能够重现DNA复制的无细胞系统，对存在DNA损伤时DNA复制的详细生化分析受到了阻碍。在这里，我们描述了基于非洲爪蟾卵提取物的实验，以研究复制叉稳定性的生化方面。
