PCR-Based Assays for the Detection and Quantitation of DNA Damage and Repair
PCR-Based Assays for the Detection and Quantitation of DNA Damage and Repair
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基于 PCR 的 DNA 损伤和修复检测和定量分析
DOI：
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发表时间：
1996
期刊：
影响因子：
--
通讯作者：
B. V. Houten
中科院分区：
文献类型：
--
作者：
F. Yakes;Yiming Chen;B. V. Houten
接触环境和内源性遗传毒性物质会导致细胞 DNA 损伤，如果这种损伤得不到修复，就会对个人造成重大健康风险。一些人类疾病，包括科凯恩综合征和色素性干皮病，都与 DNA 损伤修复缺陷有关（Friedberg 等，1995）。后者观察到 DNA 修复能力总体下降，而前者与转录偶联因子的缺陷有关，该转录偶联因子促进活性基因转录链的快速修复。分析基因和链特异性修复的一种通用方法是基于对损伤特异性核酸内切酶诱导的 DNA 链断裂的 Southern 分析（Smith 和 Mellon，1990；Bohr 和 Okumoto，1988）。这种核酸内切酶敏感位点 (ESS) 技术采用 T4 核酸内切酶 V，它在嘧啶二聚体处切割 DNA（Ganesan 等，1980），从而消除了其在碱性 Southern 分析中与放射性探针杂交的能力。尽管这种方法对于阐明单拷贝基因的基因特异性修复至关重要（Bohr et al., 1987; Mellon et al., 1987; Mellon and Hanawalt, 1989），但也存在一定的局限性。它需要一些有关感兴趣基因或基因组片段侧翼的限制序列信息、在受损碱基附近切割的病变特异性核酸内切酶，也许最重要的是需要通常用于 Southern 测定的大量 DNA (5–10 μg)。
Exposure to genotoxic agents from both environmental and endogenous sources which result in damage to cellular DNA poses a significant health risk to the individual if such damage is left unrepaired. Several human diseases, including Cockayne’s syndrome and xeroderma pigmentosum, have been associated with defects in the repair of DNA damage (Friedberg et al., 1995). An overall decrease in DNA repair capacity is observed in the latter, whereas the former is associated with a defect in a transcription-coupling factor which facilitates rapid repair in the transcribed strand of an active gene. One general method for the analysis of gene- and strand-specific repair is based on Southern analysis of DNA strand breaks induced by a damage-specific endonuclease (Smith and Mellon, 1990; Bohr and Okumoto, 1988). This endonuclease-sensitive site (ESS) technique employs the use of T4 endonuclease V which cleaves DNA at pyrimidine dimers (Ganesan et al., 1980), eliminating its ability to hybridize to a radioactive probe on alkaline Southern analysis. Although this methodology has been pivotal in the elucidation of gene-specific repair in single-copy genes (Bohr et al., 1987; Mellon et al., 1987; Mellon and Hanawalt, 1989), there are certain limitations. It requires some information regarding restriction sequence information flanking the gene or genomic segment of interest, a lesion-specific endonuclease to incise near the damaged base, and perhaps most significant is the requirement for large quantities of DNA (5–10 μg) generally used in Southern assays.
通过定量免疫偶联 PCR 评估特定基因组区域的 DNA 损伤和修复。
影响因子：
14.9
作者：
Denissenko,MF;Venkatachalam,S;Yamasaki,EF;Wani,AA
通讯作者：
Wani,AA