{"title":"Cloning and expansion of repetitive DNA sequences.","authors":"Sophie L Williams, Gideon Coster","doi":"10.1016/bs.mcb.2022.10.013","DOIUrl":null,"url":null,"abstract":"<p><p>Repeat and structure-prone DNA sequences comprise a large proportion of the human genome. The instability of these sequences has been implicated in a range of diseases, including cancers and neurodegenerative disorders. However, the mechanism of pathogenicity is poorly understood. As such, further studies on repetitive DNA are required. Cloning and maintaining repeat-containing substrates is challenging due to their inherent ability to form non-B DNA secondary structures which are refractory to DNA polymerases and prone to undergo rearrangements. Here, we describe an approach to clone and expand tandem-repeat DNA without interruptions, thereby allowing for its manipulation and subsequent investigation.</p>","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":null,"pages":null},"PeriodicalIF":0.0000,"publicationDate":"2024-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Methods in cell biology","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/bs.mcb.2022.10.013","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2022/11/28 0:00:00","PubModel":"Epub","JCR":"Q4","JCRName":"Biochemistry, Genetics and Molecular Biology","Score":null,"Total":0}
引用次数: 0
Abstract
Repeat and structure-prone DNA sequences comprise a large proportion of the human genome. The instability of these sequences has been implicated in a range of diseases, including cancers and neurodegenerative disorders. However, the mechanism of pathogenicity is poorly understood. As such, further studies on repetitive DNA are required. Cloning and maintaining repeat-containing substrates is challenging due to their inherent ability to form non-B DNA secondary structures which are refractory to DNA polymerases and prone to undergo rearrangements. Here, we describe an approach to clone and expand tandem-repeat DNA without interruptions, thereby allowing for its manipulation and subsequent investigation.
重复和结构易变的 DNA 序列在人类基因组中占很大比例。这些序列的不稳定性与一系列疾病有关，包括癌症和神经退行性疾病。然而，人们对致病机制知之甚少。因此，需要对重复 DNA 进行进一步研究。克隆和维护含重复底物具有挑战性，因为它们本身能够形成非 B 型 DNA 二级结构，而这种结构对 DNA 聚合酶具有耐受性，并且容易发生重排。在这里，我们描述了一种无中断克隆和扩增串联重复 DNA 的方法，从而可以对其进行操作和后续研究。
期刊介绍：
For over fifty years, Methods in Cell Biology has helped researchers answer the question "What method should I use to study this cell biology problem?" Edited by leaders in the field, each thematic volume provides proven, state-of-art techniques, along with relevant historical background and theory, to aid researchers in efficient design and effective implementation of experimental methodologies. Over its many years of publication, Methods in Cell Biology has built up a deep library of biological methods to study model developmental organisms, organelles and cell systems, as well as comprehensive coverage of microscopy and other analytical approaches.
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