IN VITRO ANALYSIS OF A DNA REPLICATION APPARATUS

计划使用纯化的噬菌体T4的复制蛋白， 分析在DNA过程中产生高保真度的机制 复制的 T4 DNA聚合酶的准确性（在缺乏 在复制天然DNA模板的过程中， 在许多不同的DNA序列背景下测量。 的影响 各种不同的因素，特别强调聚合酶 辅助蛋白质，对该反应的准确性将被测量。 的 这些蛋白质的作用也将通过取代突变体来研究。 7种酶催化的DNA复制反应 复制复合体 将尝试分析相对 碱基错配贡献，碱基堆积，校对效率 以及更长范围的DNA序列因素对DNA复制的准确性的影响。 将建立一个研究移码突变的体外系统， 建立了这些机制，以便能够 分析得非常详细。

It is planned to use purified replication proteins of bacteriophage T4 to analyze the mechanisms involved in generation of high fidelity during DNA replication. The accuracy of T4 DNA polymerase (in the absence of assessory proteins) during the duplication of natural DNA templates will be measured in a number of different DNA sequence contexts. The influence of a variety of different factors, with special emphasis on polymerase accessory proteins, on the accuracy of this reaction will be measured. The role of these proteins will also be investigated by substituting mutant species in DNA replication reactions catalyzed by a seven enzyme replication complex. Attempts will be made to analyze the relative contributions of base mispairing, base stacking, efficiency of proofreading and longer range DNA sequence factors on the accuracy of DNA replication. An in vitro system for the study of frameshift mutations will be established so that the mechanisms involved in their generation can be analyzed in great detail.