IONIZING RADIATION MUTAGENESIS IN HUMAN CELLS

这项研究旨在研究辐射的机制。 人类细胞中的突变。X射线可诱发不同的突变 可以通过几个类别相互区分的类别 细胞和分子技术。据推测，这些 不同的类别产生于不同类型的DNA损伤。 众所周知，辐射防护条件会影响相对产额。 一些特定的DNA损伤，因此预计它们 会改变观察到的突变的光谱。的光谱 X射线将被测定，并与所获得的光谱进行比较 在用调节产额的辐射防护剂照射后 诱变株的数量。突变谱将在以下几个方面定义 方式。细胞组件将由剂量反应数据组成。 将X连锁hprt基因座与常染色体tk基因进行比较。 发生的一类生长缓慢的突变体的测量 只有在tk才会允许更大规模的区分 基因突变导致的染色体突变。须符合的条件 研究包括单独使用X射线，或在半胱胺存在的情况下， 二甲基亚砜，或缺氧，或这些的组合 放射防护器。 光谱的分子方面将区分 Southern印迹、Northern印迹法检测基因突变类型 和DNA测序方法来比较类型和位置 HPRT基因内的突变。一组突变体将被分离出来 在单独接受X射线治疗或在各种不同的 放射防护器。所有突变体都将通过Southern杂交进行分析 确定基因突变的比例，这些基因突变是由 比例变化。每个含有HPRT DNA的突变体都将是 通过Northern印迹分析，看看它们属于四个等级中的哪一个 变成(正常的RNA，降低的水平，缩小的尺寸，没有RNA)。突变者 将克隆和测序哪些合成hprt mrna，使用 基因克隆/双脱氧测序技术。 这项研究将提供有关X射线性质的信息- 内源性人类基因的诱变。它将决定 辐射防护剂是否会对特定类别的 诱变。与已发表的关于影响的工作进行比较 辐射防护剂对DNA变化的影响将允许对 各种损伤会导致特定类别的突变。

This research is designed to study mechanisms of radiation mutagenesis in human cells. X-rays induce different mutational classes which can be distinguished from one another by several cellular and molecular techniques. It is hypothesized that these different classes arise from different types of DNA damage. Radioprotective conditions are known to affect the relative yields of certain DNA lesions, and therefore it is anticipated that they will alter the spectrum of mutations observed. The spectrum for X-rays will be determined and compared with spectra obtained following irradiation with radioprotectors which modulate the yield of induced mutants. Mutational spectra will be defined in several ways. The cellular component will consist of dose-response data. The X-linked hprt locus will be compared with the autosomal tk locus; measurement of a class of slow-growing mutants which occur only at tk will allow the differentiation of larger-scale chromosomal mutations from gene mutations. Conditions to be studied include X-rays alone, or in the presence of cysteamine, dimethylsulfoxide, or hypoxia, or combinations of these radioprotectors. The molecular aspect of the spectra will differentiate among classes of gene mutations by utilizing Southern blot, Northern blot and DNA sequencing methods to compare the types and positions of mutation within the hprt gene. Sets of mutants will be isolated after treatment with X-rays alone or in the presence of the various radioprotectors. All mutants will be analyzed by Southern blot to determine the fraction of gene mutations which arise from large- scale changes. Each mutant which contains hprt DNA will be analyzed by Northern blot to see which of four classes they fall into (normal RNA, reduced levels, reduced size, no RNA). Mutants which synthesize hprt mRNA will be cloned and sequenced, using a cDNA cloning/dideoxy sequencing technique. This research will provide information on the nature of X-ray- induced mutations in an endogenous human gene. It will determine whether radioprotectors differentially affect specific classes of induced mutations. Comparison with published work on the effects of radioprotectors on DNA change will allow an assessment of which sorts of lesions lead to particular classes of mutations.