近日，英国剑桥大学Balasubramanian, Shankar团队开发出用于检测DNA中表观遗传胞嘧啶修饰的非自然碱基对。2025年8月20日出版的《自然-化学》杂志发表了这项成果。

基因组DNA中天然的、共价修饰的胞嘧啶碱基是重要的表观遗传标记。胞嘧啶修饰的单碱基分辨率测序方法通常采用化学方法进行修饰选择性c到t代码转换，并且可能需要对复杂数据进行容易出错的减法分析。

研究组报告测序的表观遗传碱基利用一个非自然的碱基对系统。这种方法依赖于5-甲酰基胞嘧啶的丙二腈加合物和质子化的3,7-二氮杂腺嘌呤之间的氢键互补。该非自然碱基对的特异性通过生物物理DNA热熔分析和DNA聚合酶的模板定向掺入研究。通过控制3,7-二氮杂腺嘌呤的质子化状态，提高了碱基对的选择性。研究组举例说明了这种非自然碱基对的主题，在DNA模板中对5-甲酰基胞嘧啶进行测序，主题为sanger型格式。这种碱基对和一般概念在利用沃森-克里克碱基对直接识别表观遗传碱基的进一步测序平台上实施的范围是存在的。

附：英文原文

Title: An unnatural base pair for the detection of epigenetic cytosine modifications in DNA

Author: Schmidl, David, Becker, Sidney M., Edgerton, James M., Balasubramanian, Shankar

Issue&Volume: 2025-08-20

Abstract: Natural, covalently modified cytosine bases within genomic DNA function as important epigenetic markers. Approaches for single-base-resolution sequencing of cytosine modifications typically deploy chemistry for modification-selective C-to-T code conversion and can require error-prone subtractive analysis of complex data. Here we report the sequencing of an epigenetic base by exploiting an unnatural base pair system. This approach relies on hydrogen-bonding complementarity between a malononitrile adduct of 5-formylcytosine and protonated 3,7-dideazaadenine. The specificity of this unnatural base pair was studied by biophysical DNA thermal melting analysis and by template-directed incorporation by DNA polymerase enzymes. Base pair selectivity was enhanced by controlling the protonation state of 3,7-dideazaadenine. We exemplify use of this unnatural base pair to sequence 5-formylcytosine in a DNA template using a Sanger-type format. There is scope for this base pair and the general concept to be implemented on further sequencing platforms that exploit Watson–Crick base pairing to directly identify epigenetic bases.

DOI: 10.1038/s41557-025-01925-6

Source: https://www.nature.com/articles/s41557-025-01925-6