DETECTION AND MEASUREMENT OF HUMAN DNA ADDUCTS

人类 DNA 加合物的检测和测量

• 批准号: 3548870
• 负责人: KURT RANDERATH
• 金额: $ 16.36万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1986
• 资助国家: 美国
• 起止时间: 1986-08-01 至 1990-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3548870
• 关键词: DNA; aging; autoradiography; azo compounds; carbopolycyclic compound; chemical carcinogen; chemical fingerprinting; dyes; environment related neoplasm /cancer; environmental toxicology; formaldehyde; gas poisoning; laboratory mouse; laboratory rat; occupational hazard; polystyrenes; radioassay; radiotracer; thin layer chromatography; tobacco abuse; DNA; aging; autoradiography; azo compounds; carbopolycyclic compound; chemical carcinogen; chemical fingerprinting; dyes; environment related neoplasm /cancer; environmental toxicology; formaldehyde; gas poisoning; laboratory mouse; laboratory rat; occupational hazard; polystyrenes; radioassay; radiotracer; thin layer chromatography; tobacco abuse

A 32P-Postlabeling method developed in the applicant's laboratory will be applied to study potential covalent DNA lesions in human white blood cells and tissues exposed to occupational or environmental mixtures of genotoxic compounds. The basic procedure entails the enzymatic digestion of the DNA preparation to be tested for the presence of adducts to deoxyribonucleoside 3'-monophosphates, conversion of these mononucleotides to 5'-32P-labeled deoxyribonucleoside 3',5'-bisphosphates, resolution of the 32P-labeled nucleotides by thin-layer chromatography, and autoradiography. 32P-Labeled adduct fractions are detected as extra spots on X-ray film and are quantitated by scintillation (Cerenkov) counting. Characteristic "fingerprints" are obtained for DNA adducts formed in vivo from authentic carcinogens or mixtures of genotoxic compounds such as cigarette smoke. The 32P-postlabeling assay will be suitable specifically for the analysis of traces of adducts in microgram amounts of human DNA. For the characterization of adducts observed in human DNA, co-chromatography and stability studies will be conducted with adducts obtained from DNA of experimental animals treated with crude environmental/occupational mixtures or authentic carcinogens known to occur in the exposure of interest. Cigarette smoking will be considered as a confounding variable on the basis of the detection of specific smoking-associated DNA adducts. The project will focus on occupational exposures to polycyclic aromatic hydrocarbons, aromatic amines/amides, azo compounds, dyestuffs, formaldehyde, and styrene. In addition, it is proposed to study whether the exposure to emissions of wood-burning stoves and the natural process of aging give rise to adducts in human DNA.

申请人实验室中开发的32p固定方法将是 应用于研究人类白细胞中潜在的共价DNA病变 以及暴露于遗传毒性的职业或环境混合物的组织 化合物。 基本过程需要DNA的酶促消化 准备对脱氧核苷的加合物进行测试 3'-onophophates，这些单核苷酸转化为5'-32p标记的 脱氧核苷3'，5'生磷酸盐，32p标记的分辨率 通过薄层色谱和放射自显影的核苷酸。 32p标记 在X射线膜上检测到加合物的分数是 通过闪烁（Cerenkov）计数定量。 特征 对于由真实的体内形成的DNA加合物获得的“指纹” 遗传毒性化合物（例如香烟烟雾）的致癌物或混合物。 32p固定标签测定法将是专门用于分析的 微克量的人DNA中的加合物的痕迹。 为了表征人DNA中观察到的加合物 将与加合物一起进行共染色和稳定研究 从用粗糙处理的实验动物的DNA获得 已知发生的环境/职业混合物或正宗的致癌物 在兴趣的情况下。 吸烟将被视为 根据特定的检测而混淆变量 吸烟相关的DNA加合物。 该项目将集中于对多环芳香族的职业暴露 碳氢化合物，芳香胺/酰胺，偶氮化合物，染料，染料， 甲醛和苯乙烯。 此外，建议研究是否 暴露于燃木炉的排放和自然过程 衰老会引起人DNA的加合物。