Haseltine W A, Franklin W, Lippke J A
Environ Health Perspect. 1983 Feb;48:29-41. doi: 10.1289/ehp.834829.
The use of a postlabeling method to characterize and to detect infrequent base modifications in DNA is outlined. This method has the advantage that low levels of DNA modifications, approximately 1 modified base per 10(5) nucleotides, can be detected. Moreover, a broad spectrum of modification can be identified by using this methodology. The basis for the method involves transfer of a radioactive phosphate from the gamma position of ATP to the 5'-hydroxyl terminus of 3'-phosphoryl nucleotides that are derived from modified DNA by appropriate nuclease digestion. The second method involves use of a defined DNA sequence within human cells. The alpha sequence is used as a probe for DNA damage to specific nucleotides. The alpha DNA sequence is reiterated approximately 300,000 times in the human genome and exists in tandem arrays. It comprises approximately 1% of the entire genome. The reiterated sequence is sufficiently homogeneous to permit its use as a probe for a site specific in DNA damage. Examples of the application of both of these methodologies to DNA damage inflicted in human cells by chemicals and ultraviolet light are provided.
概述了一种用于表征和检测DNA中罕见碱基修饰的标记后方法。该方法的优点是能够检测低水平的DNA修饰，大约每10⁵个核苷酸中有1个修饰碱基。此外，使用这种方法可以鉴定广泛的修饰类型。该方法的基础是通过适当的核酸酶消化，将放射性磷酸从ATP的γ位转移到由修饰DNA衍生的3'-磷酸核苷酸的5'-羟基末端。第二种方法涉及在人类细胞中使用特定的DNA序列。α序列用作检测特定核苷酸DNA损伤的探针。α DNA序列在人类基因组中大约重复300,000次，并以串联阵列形式存在。它约占整个基因组的1%。该重复序列足够均匀，可作为DNA损伤特定位点的探针。提供了这两种方法在化学物质和紫外线对人类细胞造成的DNA损伤中的应用实例。
