通过工程性二氧化酶NTET辅助eNAPS直接测序DNA5-甲基细胞蛋白
Shan Zhang1, Neng-Bin Xie1, Li Zeng1
1Department of Occupational and Environmental Health, School of Public Health, Department of Radiation and Medical Oncology, Zhongnan Hospital of Wuhan University, State Key Laboratory of Metabolism and Regulation in Complex Organisms, Wuhan University Wuhan 430071 China bfyuan@whu.edu.cn jxiong@whu.edu.cn.
Chemical science
|July 21, 2025
概括
我们开发了工程NTET辅助胺基测序 (eNAPS) 技术,以在单基分辨率下准确检测DNA甲基化 (5-甲基素,5mC). 这种新的方法为表观遗传研究和疾病研究提供了更好的灵敏度和效率.
科学领域:
- 表观遗传学和分子生物学
- 基因组学和生物信息学
- 癌症研究 癌症研究
背景情况:
- DNA甲基化 (5-甲基细胞素,5mC) 是哺乳动物的一个关键的表观遗传修饰,涉及到疾病发病和癌症.
- 精确的单基分辨率检测5mC对于理解其生物作用至关重要,但目前的方法在分辨率,偏差和准确性方面存在局限性.
- 现有的二硫酸盐测序 (BS-seq) 方法具有破坏性,并可能引入不准确性.
研究的目的:
- 开发一种新的,高度准确和高效的方法,用于定量,单基分辨率检测DNA甲基化 (5mC).
- 为改进5mC检测能力设计一种改进的TET类蛋白质.
- 为了克服5mC分析的传统二硫酸盐测序的局限性.
主要方法:
- 将Naegleria的TET样蛋白 (NTET) 改造成具有增强氧化活性和5mC的序列兼容性的重组工程NTET (eNTET).
- 通过将 eNTET 氧化与降解相结合,开发了工程 NTET 辅助的胺序列 (eNAPS).
- eNAPS将5mC氧化为5-formylcytosine (5fC) 和5-carboxylcytosine (5caC),然后在测序过程中还原为二甲 (DHU) 并读取为丁氨酸 (T).
主要成果:
- eNAPS可直接将5mC转化为T,从而在单基分辨率下准确地绘制DNA甲基化.
- 与BS-seq相比,eNAPS显示了非破坏性特性,提高了灵敏度,提高了准确性和更高的效率.
- 使用eNAPS,对肺瘤和相邻正常组织的基因组DNA成功进行了5mC的定量分析.
结论:
- eNAPS是一种温和的,不含二硫酸盐的,高精度的方法,用于在单基分辨率下定量检测5mC.
- 这种技术为研究5mC在表观遗传调节中的动态性质提供了有价值的工具.
- eNAPS有助于更深入地了解5mC在疾病发病的作用,特别是癌症的发展.
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