...
  • 主题
高级搜索  >
历史搜索 清空历史
首页> 外文期刊>Nucleic acids research >Detection of DNA–protein crosslinks (DPCs) by novel direct fluorescence labeling methods: distinct stabilities of aldehyde and radiation-induced DPCs
【24h】

Detection of DNA–protein crosslinks (DPCs) by novel direct fluorescence labeling methods: distinct stabilities of aldehyde and radiation-induced DPCs

机译：通过新颖的直接荧光标记方法检测DNA-蛋白质交联（DPC）：醛和辐射诱导的DPC具有独特的稳定性

获取原文
   

获取外文期刊封面封底 >>

       
页面导航
  • 摘要
  • 著录项
  • 相似文献
  • 相关主题

摘要

Proteins are covalently trapped on DNA to form DNA–protein crosslinks (DPCs) when cells are exposed to DNA-damaging agents. DPCs interfere with many aspects of DNA transactions. The current DPC detection methods indirectly measure crosslinked proteins (CLPs) through DNA tethered to proteins. However, a major drawback of such methods is the non-linear relationship between the amounts of DNA and CLPs, which makes quantitative data interpretation difficult. Here we developed novel methods of DPC detection based on direct CLP measurement, whereby CLPs in DNA isolated from cells are labeled with fluorescein isothiocyanate (FITC) and quantified by fluorometry or western blotting using anti-FITC antibodies. Both formats successfully monitored the induction and elimination of DPCs in cultured cells exposed to aldehydes and mouse tumors exposed to ionizing radiation (carbon-ion beams). The fluorometric and western blotting formats require 30 and 0.3?μg of DNA, respectively. Analyses of the isolated genomic DPCs revealed that both aldehydes and ionizing radiation produce two types of DPC with distinct stabilities. The stable components of aldehyde-induced DPCs have half-lives of up to days. Interestingly, that of radiation-induced DPCs has an infinite half-life, suggesting that the stable DPC component exerts a profound effect on DNA transactions over many cell cycles.
机译：当细胞暴露于DNA破坏剂时，蛋白质被共价捕获在DNA上，形成DNA-蛋白质交联（DPC）。 DPC会干扰DNA交易的许多方面。当前的DPC检测方法通过与蛋白质相连的DNA间接测量交联蛋白质（CLP）。但是，这种方法的主要缺点是DNA和CLP数量之间存在非线性关系，这使得定量数据解释变得困难。在这里，我们开发了基于直接CLP测量的DPC检测的新方法，其中，用异硫氰酸荧光素（FITC）标记从细胞中分离的DNA中的CLP，并通过荧光法或使用抗FITC抗体的蛋白质印迹法进行定量。两种格式均成功监测了暴露于醛类的培养细胞和暴露于电离辐射（碳离子束）的小鼠肿瘤中DPC的诱导和消除。荧光印迹法和蛋白质印迹法分别需要30和0.3？μgDNA。对分离的基因组DPC的分析表明，醛类和电离辐射均会产生两种类型的具有不同稳定性的DPC。醛诱导的DPC的稳定成分的半衰期可长达数天。有趣的是，辐射诱导的DPC具有无限的半衰期，这表明稳定的DPC成分在许多细胞周期中对DNA交易产生了深远的影响。

著录项

相似文献

  • 外文文献
  • 中文文献
  • 专利
获取原文

客服邮箱：kefu@zhangqiaokeyan.com

京公网安备：11010802029741号 ICP备案号：京ICP备15016152号-6 六维联合信息科技 (北京) 有限公司©版权所有

  • 客服微信

  • 服务号