X-RAY DAMAGE AND REPAIR OF PRIMATE CELL A DNA SEQUENCES

电离后特定DNA碱基序列损伤与修复的知识 辐射可以为辐射的分子机制提供线索 损伤会导致突变、致癌和细胞周期紊乱 与放射治疗相关。我们现在可以得到这样的信息 因为DNA碱基测序方法的进步。这些方法是 易于应用于研究同质DNA群体，如 噬菌体DNA和灵长类DNA的高重复组分α(172 B.P.重复)。阿尔菲脱氧核糖核酸存在于所有人类细胞中；它们解释了 25%的非洲绿猴细胞DNA及其碱基序列是已知的。 来自连续系BSC-1和CV-1的猴细胞将受到照射和 利用EcoR1*、Hind III和Hae 111分离到它们的α序列 细菌、琼脂糖凝胶和聚丙烯酰胺凝胶电泳法 分离。经过5英尺32P磷酸盐末端标记和二次 限制，链断裂的位置将由MAXAM和 吉尔伯特碱基测序法。目前还没有关于 电离辐射对活细胞DNA碱基序列的破坏。 尽管如此，学习这一点的技术是可用的， 实验是可行的。对纯化的DNA进行了体外辐射研究 最近发表的；一些初步的碱基序列损伤已经 特色化的。我们建议了解特定的碱基序列变化 它们与活细胞相关。 X-射线剂量反应与碱基体内修复的时间进程 将研究顺序损坏，并将其与 缺氧状态。同步的HeLa和猴子细胞将被照射 以及通过流式细胞仪监测它们在整个周期中的进展。 辐射后未被纠正的预突变碱基序列变化 现在可以第一次被识别。 将研究咖啡因促进2人逃脱2人被捕 和碱基序列损伤修复的进展。 咖啡因的存在和缺失。比较研究与穿插 将产生重复的灵长类序列，Kpn1 1.2kb家族，因为 这些新发现的重复序列分布广泛；它们 被转录，它们的碱基序列和位置与 特定的人类致癌基因正在研究中。

Knowledge of specific DNA base sequence damage and repair after ionizing radiation can provide clues to the molecular mechanisms by which radiation damage leads to mutation, carcinogenesis, and cell cycle disturbances relevant to radiation therapy. We can now get this kind of information because of advances in DNA base sequencing methods. The methods are readily applied to study of homogeneous DNA populations such as bacteriophage DNA and highly repetitive component Alpha of primate DNA (172 b.p. repeat). Alphoid DNA's are found in all human cells; they account for 25% of African green monkey cell DNA and their base sequences are known. Monkey cells from continuous lines BSC-1 and CV-1 will be irradiated and their Alpha sequences isolated using EcoR1*, Hind III, and Hae 111 endonuleases and agarose and polyacrylamide gel electrophoretic separation. After 5 feet 32p phosphate end-labeling and secondary restriction, the sites of strand scissions will be studied by the Maxam and Gilbert base sequencing method. There is no information yet available on DNA base sequence damage by ionizing radiation in living cells. Neverthesless, the techniques to learn this are available and the experiments are feasible. In vitro radiation studies on purified DNA were recently published; some preliminary base sequence damage has already been characterized. We proposed to learn the specific base sequence changes which are relevant in living cells. X-ray does-responses and the time course of in vivo repair of the base sequence damage will be studied, comparing this with damage induced under anoxic conditions. Synchronized HeLa and monkey cells will be irradiated and their progress through the cycle monitored by flow cytometry. Premutational base sequence changes which are not corrected after radiation can now be identified for the first time. Studies will be made on caffeine promoted escape from 2 arrest in irradiated cells, and on the progress of base sequence damage repair in the presence and absence of caffeine. Comparative studies with interspersed repetitive primate sequences, the Kpnl 1.2kb family, will be made because these newly discovered repetitive sequences are widely distributed; they are transcribed, and their base sequences and location in relation to specific human oncogenes are under study.