Department of Health Sciences and Technology , ETH Zürich , Schmelzbergstrasse 9 , 8092 Zürich , Switzerland.
J Am Chem Soc. 2018 Aug 8;140(31):9783-9787. doi: 10.1021/jacs.8b03715. Epub 2018 Jul 26.
Single-nucleotide-resolution sequencing of DNA damage is required to decipher the complex causal link between the identity and location of DNA adducts and their biological impact. However, the low abundance and inability to specifically amplify DNA damage hinders single-nucleotide mapping of adducts within whole genomes. Despite the high biological relevance of guanine oxidation and seminal recent advances in sequencing bulky adducts, single-nucleotide-resolution whole genome mapping of oxidative damage is not yet realized. We coupled the specificity of repair enzymes with the efficiency of a click DNA ligation reaction to insert a biocompatible locator code, enabling high-throughput, nucleotide-resolution sequencing of oxidative DNA damage in a genome. We uncovered thousands of oxidation sites with distinct patterns related to transcription, chromatin architecture, and chemical oxidation potential. Click-code-seq overcomes barriers to DNA damage sequencing and provides a new approach for generating comprehensive, sequence-specific information about chemical modification patterns in whole genomes.
要破解 DNA 加合物的身份和位置与其生物学影响之间复杂的因果关系，就需要对 DNA 损伤进行单核苷酸分辨率测序。然而，DNA 损伤的低丰度和无法特异性扩增阻碍了整个基因组中加合物的单核苷酸图谱绘制。尽管鸟嘌呤氧化具有很高的生物学相关性，并且最近在测序大体积加合物方面取得了重大进展，但单核苷酸分辨率的全基因组氧化损伤图谱绘制尚未实现。我们将修复酶的特异性与点击 DNA 连接反应的效率相结合，插入了一个生物相容性定位码，从而能够在基因组中进行高通量、核苷酸分辨率的氧化 DNA 损伤测序。我们发现了数千个与转录、染色质结构和化学氧化电位相关的具有独特模式的氧化位点。Click-code-seq 克服了 DNA 损伤测序的障碍，为生成整个基因组中化学修饰模式的全面、序列特异性信息提供了一种新方法。
