Genetic Assays to Study Repeat Fragility in Saccharomyces cerevisiae
Genetic Assays to Study Repeat Fragility in Saccharomyces cerevisiae
复制标题
研究酿酒酵母重复脆弱性的遗传分析
DOI：
10.1007/978-1-4939-9784-8_5
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发表时间：
2019
影响因子：
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通讯作者：
Freudenreich, Catherine H
中科院分区：
文献类型：
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作者：
Polleys, Erica J.;Freudenreich, Catherine H
三核苷酸重复序列在人类基因组中很常见，并且可以经历重复数量的变化并导致长度依赖性染色体脆性。扩增的CAG重复序列与超过14种人类疾病有关，并且被认为是断裂和基因组重排的热点。在这里，我们描述了两种基于酵母的检测方法，用于评估重复序列中发生的染色体断裂率（脆性），并通过变异来评估转录的作用。第一脆性测定利用末端丢失和随后的端粒添加作为酵母人工染色体（YAC）的主要修复模式。第二种脆性测定依赖于染色体断裂刺激重组介导的修复的事实。基于PCR的测定可用于在用于测量重复脆性的相同条件下评价重复的不稳定性。这些检测有助于了解导致染色体断裂和不稳定三核苷酸重复序列长度变化的遗传机制。
Trinucleotide repeats are common in the human genome and can undergo changes in repeat number and cause length-dependent chromosome fragility. Expanded CAG repeats have been linked to over 14 human diseases and are considered hotspots for breakage and genomic rearrangement. Here we describe twoSaccharomyces cerevisiaebased assays that evaluate the rate of chromosome breakage that occurs within a repeat tract (fragility), with variations that allow the role of transcription to be evaluated. The first fragility assay utilizes end-loss and subsequent telomere addition as the main mode of repair of a yeast artificial chromosome (YAC). The second fragility assay relies on the fact that a chromosomal break stimulates recombination-mediated repair. A PCR-based assay can be used to evaluate instability of the repeat in the same conditions used to measure repeat fragility. These assays have contributed to understanding the genetic mechanisms that cause chromosome breaks and tract-length changes at unstable trinucleotide repeats.