Volkert Michael R, Wang Jen-Yeu, Elliott Nathan A
Department of Molecular Genetics and Microbiology, University of Massachusetts Medical School, Worcester, MA, USA.
Methods Mol Biol. 2008;477:331-42. doi: 10.1007/978-1-60327-517-0_25.
In order to develop a more complete understanding of the genes required for resistance to oxidative DNA damage, we devised methods to identify genes that can prevent or repair oxidative DNA damage. These methods use the oxidative mutator phenotype of a repair deficient E. coli strain to measure the antimutator effect resulting from the expression of human cDNAs. The method can be adapted to characterize the function, and to determine the active site domains, of putative antimutator genes. Since bacteria do not contain subcellular compartments, genes that function in mitochondria, the cytoplasm, or the nucleus can be identified. Methods to determine the localization of genes in their normal host organism are also described.
为了更全面地了解抵抗氧化性DNA损伤所需的基因,我们设计了一些方法来鉴定能够预防或修复氧化性DNA损伤的基因。这些方法利用修复缺陷型大肠杆菌菌株的氧化性突变体表型来测量人cDNA表达产生的抗突变效应。该方法可用于表征假定抗突变基因的功能,并确定其活性位点结构域。由于细菌不含有亚细胞区室,因此可以鉴定在线粒体、细胞质或细胞核中发挥作用的基因。本文还描述了确定基因在其正常宿主生物体中定位的方法。