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通过超高性能液体色谱学与补充质谱学相结合,对RNA修饰的高通量分析:方法,质量控制和应用

Gefei Huang¹, Feng Zhang¹, Dongying Xie²

¹State Key Laboratory of Environmental and Biological Analysis, Department of Chemistry, Hong Kong Baptist University, Hong Kong, 999077, China.

|June 1, 2023

概括

一个新的超高性能液态染色体质谱平台在不到16分钟的时间内准确识别和量化64种RNA修饰,推进生物标志物发现和RNA生物学研究.

科学领域:

生物化学生物化学
分析化学分析化学
分子生物学分子生物学

背景情况:

下一代测序在准确表征和量化各种RNA修饰方面存在局限性.
挑战包括缺乏适合的转化反应和针对众多RNA修饰的特定抗体.
跨发育阶段和细胞类型的RNA修饰变异进一步复杂化了分析.

研究的目的:

开发一个高通量分析平台,用于全面的RNA修饰分析.
在合成和生物样本中识别和量化广泛的RNA修饰.
建立一种可靠的方法来研究模型生物中的RNA修饰景观.

主要方法:

结合超高性能液体色谱仪 (UPLC) 与补充质谱仪 (MS).
使用监控系统开发两种独立的RNA纯度交叉验证方法.
在合成样本和三个模型生物中的RNA修饰的分析:C. elegans,E. coli和A. thaliana.

主要成果:

在单个16分钟的UPLC-MS运行中,成功监测了64种类型的RNA修饰,包括定位异构体和超变 ribonucleosides.
在C. elegans,大肠杆菌和A. thaliana.之间证明了改造的核糖核酸的身份和度的显著变化.
验证了平台的实用性,用于研究响应环境线索的RNA修饰动态,并识别RNA修饰酶.

结论:

开发的UPLC-MS平台为分析RNA修饰提供了一种全新的,全面的方法.
这种方法有利于生物标志物发现的分析化学家和RNA修饰功能研究的生物学家.
该平台为探索不同物种和条件的复杂RNA修饰世界提供了强大的工具.

关键词:

凯诺哈比迪斯的优雅的植物.质量控制质量控制通过RNA甲基化.基因组RNA的修改基因组RNA的改变里核酸是一种核酸.

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