无亚硫酸盐和无减法的 5-甲基胞嘧啶及其氧化衍生物的碱基分辨率特异性测序。

Subtraction-free and bisulfite-free specific sequencing of 5-methylcytosine and its oxidized derivatives at base resolution.

机构信息

Ludwig Institute for Cancer Research, Nuffield Department of Medicine, University of Oxford, Oxford, OX3 7FZ, UK.

Target Discovery Institute, Nuffield Department of Medicine, University of Oxford, Oxford, OX3 7FZ, UK.

出版信息

Nat Commun. 2021 Jan 27;12(1):618. doi: 10.1038/s41467-021-20920-2.

Abstract

Although various methods have been developed for sequencing cytosine modifications, it is still challenging for specific and quantitative sequencing of individual modification at base-resolution. For example, to obtain both true 5-methylcytosine (5mC) and true 5-hydroxymethylcytosine (5hmC) information, the two major epigenetic modifications, it usually requires subtraction of two methods, which increases noise and requires high sequencing depth. Recently, we developed TET-assisted pyridine borane sequencing (TAPS) for bisulfite-free direct sequencing of 5mC and 5hmC. Here we demonstrate that two sister methods, TAPSβ and chemical-assisted pyridine borane sequencing (CAPS), can be effectively used for subtraction-free and specific whole-genome sequencing of 5mC and 5hmC, respectively. We also demonstrate pyridine borane sequencing (PS) for whole-genome profiling of 5-formylcytosine and 5-carboxylcytosine, the further oxidized derivatives of 5mC and 5hmC. This work completes the set of versatile borane reduction chemistry-based methods as a comprehensive toolkit for direct and quantitative sequencing of all four cytosine epigenetic modifications.

摘要

虽然已经开发出了各种方法来对胞嘧啶修饰进行测序,但要实现单个修饰在碱基分辨率上的特异性和定量测序仍然具有挑战性。例如,为了获得两种主要的表观遗传修饰(5-甲基胞嘧啶(5mC)和 5-羟甲基胞嘧啶(5hmC))的真实信息,通常需要两种方法相减,这会增加噪声并需要高测序深度。最近,我们开发了 TET 辅助的吡啶硼烷测序(TAPS)方法,用于无亚硫酸氢盐的直接测序 5mC 和 5hmC。在这里,我们证明了两种姐妹方法,TAPSβ 和化学辅助的吡啶硼烷测序(CAPS),可分别有效地用于无减法和特异性的全基因组测序 5mC 和 5hmC。我们还展示了吡啶硼烷测序(PS)用于 5-甲酰基胞嘧啶和 5-羧基胞嘧啶(5mC 和 5hmC 的进一步氧化衍生物)的全基因组分析。这项工作完成了一套基于多功能硼烷还原化学的方法,作为直接和定量测序所有四种胞嘧啶表观遗传修饰的综合工具包。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/a022/7840749/3d37ebe0f998/41467_2021_20920_Fig1_HTML.jpg

文献AI研究员

20分钟写一篇综述,助力文献阅读效率提升50倍

立即体验

用中文搜PubMed

大模型驱动的PubMed中文搜索引擎

马上搜索