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ANALYSIS OF LARGE DNA PALINDROME FORMATION IN YEAST

酵母中大 DNA 回文结构的分析

基本信息

批准号：
6028228
负责人：
DAVID Kimon BUTLER
金额：
$ 8.59万
依托单位：
MONTANA STATE UNIVERSITY -BILLINGS
依托单位国家：
美国
项目类别：
财政年份：
2000
资助国家：
美国
起止时间：
2000-03-01 至 2004-02-29
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/6028228
关键词：
DNA repair Saccharomyces cerevisiae fungal genetics gene mutation genetic recombination nucleic acid sequence plasmids

项目摘要

Selective gene amplification is a widespread phenomenon. Many amplified genes, including oncogenes in tumor cells and many genes of lower eukaryotes, are organized as part of large inverted duplications, or palindromes. Such palindromic sequences are generated from non- palindromic precursors as an initial, and presumably critical, step in gene amplification. Very little is known about how large palindromic DNA is formed. Previous experiments have established that the yeast, Saccharomyces cerevisiae, is an excellent model system for studying palindrome formation. In yeast a linear DNA strand break is introduced next to a 42 base pair (bp) inverted repeat sequence. Conversion of the broken plasmid to a palindrome is strictly dependent on the 42 bp inverted repeats. A mutation in RAD52, a gene required for repair of DNA double-strand breaks (DSBs) by homologous recombination, blocks palindrome formation. The objective of this proposal is to continue the molecular and genetic analysis of DNA palindrome formation in Saccharomyces cerevisiae. The first experiments will address whether palindromic DNA forms by an inter-molecular recombination reaction (that is, a reaction between two identical fragments in a head-to-head arrangement) or by an unusual intra-molecular recombination reaction, as it apparently does in another well studied examined of palindrome formation (see below). The second set of experiments will explore the connections between DSB repair and palindrome formation. Specifically, the effect on palindrome formation of mutations in a variety of DSB repair genes will be tested. The results of these experiments will yield valuable insights into the molecular mechanism and genetic control of this widespread and fascinating type of genome rearrangement.

基因选择性扩增是一种普遍存在的现象。许多扩增的基因，包括肿瘤细胞中的癌基因和许多低等真核生物的基因，被组织成大型反向复制或回文的一部分。这样的回文序列是由非回文前体产生的，作为基因扩增的初始步骤，可能也是关键步骤。关于回文DNA是如何形成的，我们知之甚少。以前的实验已经证明，酵母是研究回文形成的一个很好的模型系统。在酵母中，在42个碱基对(BP)的反向重复序列旁边引入了线性DNA链断裂。断裂的质粒到回文的转换严格依赖于42个碱基的反向重复序列。RAD52是一种通过同源重组修复DNA双链断裂(DSB)所需的基因，它的突变阻止了回文的形成。这项建议的目的是继续对酿酒酵母DNA回文形成的分子和遗传分析。第一个实验将解决回文DNA是通过分子间重组反应(即两个相同片段之间的反应，以头对头的方式排列)还是通过不寻常的分子内重组反应形成的，就像在另一个研究很好的回文形成过程中所做的那样(见下文)。第二组实验将探索DSB修复和回文形成之间的联系。具体地说，将测试各种DSB修复基因突变对回文形成的影响。这些实验的结果将为这种广泛而迷人的基因组重排的分子机制和基因控制提供有价值的见解。