MECHANISM FOR GENETIC RECOMBINATION IN MAMMALIAN CELLS

有丝分裂哺乳动物细胞的重组机制 催化了许多不同类型的DNA重排， 改变基因的表达或它们的空间结构。 直到 最近，这些事件很难分析，因为合适的 遗传学和模型系统还不能用来探测这些多个 重组活动。 因此，我们对 重组的机制或酶学，即使这些过程 可能对大多数细胞的正常运作很重要。 我们解决这个问题的方法是使用简单的模型系统， 体外构建重组底物，以分析不同的 重组事件 这些底物是真核表达载体 含有显性选择标记的截短或突变拷贝 基因，这样相互重组或基因转换恢复了一个 在短暂感染或稳定感染后， 将这些底物DNA整合到合适的细胞中。 我们有兴趣研究的频率和机制， 染色体外、染色体内和染色体间、相互和 非相互重组 因为染色体外重组 新转染的DNA似乎比染色体上的DNA更常见。 重组，我们希望确定这些不同的基础 频率. 缺口修复及其可能的应用进行了探讨。 我们长期利益之一是遗传信息交流 染色体和染色体外质粒之间，以及 重组的酶学。 因此，我们计划探索方法， 基因靶向广告基因恢复，并希望开发一种体外系统， 研究重组。 许多互惠和非互惠的assasy 事件也将扩展到人类细胞，包括那些 被认为是异常的重组和修复，和影响， 药物和DNA损伤剂将被评估。 我们希望， 这些基本过程将有助于更好地理解 重组发生以及它如何在哺乳动物细胞中被调节。

Mitotically dividing mammalian cells posses the recombinational machinery to catalyze a number of different types of DNA rearrangements that can alter either the expression of genes or their spatial configuration. Until recently, these events have been difficult to analyze because suitable genetic and model systems have not been available to probe these multiple recombinational activities. As a consequence, very little is known about the mechanisms or enzymology of recombination, even though these processes are likely to be important for the proper functioning of most cells. Our approach to this problem has been to use simple model systems, based on recombination substrates constructed in vitro to analyze different recombinatin events. These substrates are eukaryotic expression vectors containing either truncated or mutant copies of dominant selectable marker genes, such that reciprocal recombination or gene conversion restores a functiinal copy of the gene, following transient infection or stable integration of these substrate DNAs into appropriate cells. We are interested in studying the frequencies and mechanisms of extrachromosomal, intrachromosomal and interchromosomal, reciprocal and non-reciprocal recombination. Because extrachromosomal recombination of newly transfected DNA appears to be much more frequent than chromosomal recombination, we wish to determine the basis for these different frequencies. Gap repair and its possible applicatious will be explored. Among our long term interests are the exchange of genetic informatin between the chromosome and extrachromosomal plasmids, as well as the enzymology of recombination. We therefore, plan to explore methods for gene targeting ad gene recovery and hope to develop an in vitro system to study recombination. Many of the assasy for reciprocal and non-reciprocal events will also be extended to human cells including those that are believed to be aberrant in recombination and repair, and the effect of drugs and DNA damaging agents will be evaluated. We hope that the study of these basic processes will contribute to a better understanding of how recombination occurs and how it might be modulated in mammalian cells.