化学学报 ›› 2013, Vol. 71 ›› Issue (01): 26-35.DOI: 10.6023/A12110891 上一篇    下一篇

综述

DNA羟甲基化测序技术

赵超, 汪海林   

  1. 中国科学院生态环境研究中心 环境化学与生态毒理学国家重点实验室 北京 100085
  • 收稿日期:2012-11-09 出版日期:2013-01-14 发布日期:2012-12-25
  • 通讯作者: 汪海林 E-mail:hlwang@rcees.ac.cn
  • 基金资助:

    项目受科技部(Nos. 2009CB421605, 2010CB933502, 2011YQ060084)和国家自然科学基金(Nos. 21077129, 20921063, 20877091, 20890112, 21125523)资助

The Progress on Sequencing and Detection of Hydroxymethylated DNA

Zhao Chao, Wang Hailin   

  1. State Key Laboratory of Environmental Chemistry and Ecotoxicology, Research Center for Eco-environmental Sciences, Chinese Academy of Sciences, Beijing 100085
  • Received:2012-11-09 Online:2013-01-14 Published:2012-12-25
  • Supported by:

    Project supported by the National Basic Research Program of China (Nos. 2009CB421605, 2010CB933502 and 2011YQ060084) and the National Natural Science Foundation of China (Nos. 21077129, 20921063, 20877091, 20890112, and 21125523).

5-羟甲基胞嘧啶(5-hydroxymethylcytosine, 5hmC)是继5-甲基胞嘧啶(5-methylcytosine, 5mC)之后发现的第六种碱基, 已在多种哺乳动物组织和细胞中检出. 与5mC相比, 5hmC的含量更低, 但5hmC也具有非常重要的生物学功能. 已有的研究表明, 5hmC参与了染色体重新编程、基因表达的转录调控, 并在DNA去甲基化过程中发挥重要作用. 另外, 5hmC可能与特定肿瘤的发生密切相关, 有可能成为肿瘤早期诊断的生物标志物. 因此, 发展可靠、灵敏和准确的5hmC检测技术至关重要. 本综述针对DNA羟甲基化的检测及测序技术进行了简要介绍.

关键词: DNA羟甲基化, DNA甲基化, DNA测序技术

The recently re-discovered 5-hydroxymethylcytosine (5hmC) is established as the sixth base and found in various mammalian tissues. Although the content of 5hmC is much lower than that of the best known epigenetic mark 5-methylcytosine (5mC), 5hmC plays key roles in nuclear reprogramming, regulates the gene activity, and initiates the DNA demethylation in mammals. The formation of 5hmC results from the oxidation of 5mC in genomic DNA, which is mediated by Tet (ten eleven translocation) family dioxygenases. The Tet-mediated 5mC oxidation has just been identified. The 5hmC formation can cause DNA replication-dependent and passive DNA demethylation, meanwhile, it can also induce active DNA demethylation by further oxidation forming 5-formylcytosine (5fC) and 5-carboxylcytosine (5caC), which can be removed by base excision repair. Essentially, the discovery of 5fC and 5caC indicates the occurrence of active DNA demethylation mechanisms. The 5fC and 5caC have been detected in genomic DNA of mouse embryonic stem cells, but 5hmC has been found in various tissues. The 5hmC abundance in genomic DNA of certain tumors is found to be associated with the tumor process. However, the genome-wide distribution and sequence selectivity of 5hmC are not known yet. 5hmC cannot be discriminated from 5mC by the well-known bisulfite sequencing. Therefore, it is essential to develop new methods and technologies for detecting and sequencing 5hmC. To this purpose, the two or more of the technologies for modification of 5hmC by glucosylation or chemical labeling, restriction enzymatic digestion, affinity capture, liquid chromatography and mass spectrometry, and bisulfite sequencing are combined. The further combination with next-generation high-throughput DNA sequencing technologies may provide genome-wide information. The single molecule real time DNA sequencing is also of choice to detect 5hmC at the gene level or the genome level. Recently, oxidative bisulfite sequencing was for the first time developed for quantitative mapping of 5hmC in genomic DNA at single base resolution. The genome-wide 5hmC sequencing at single base resolution is also reported. Here we briefly review and discuss the detection and sequencing technologies for 5hmC analysis.

Key words: 5-hydroxymethylcytosine, 5-methylcytosine, DNA sequencing