核糖核酸
化学
胞嘧啶
核苷酸
转录组
RNA序列
计算生物学
DNA
DNA测序
过渡(遗传学)
基因
基因表达
生物化学
生物
作者
Li Su,Feng Chen,Huahang Yu,Hao Yan,Fengjiao Zhao,Chunhai Fan,Yongxi Zhao
标识
DOI:10.1021/acs.analchem.1c03361
摘要
Dynamic information of intracellular transcripts is essential to understand their functional roles. Routine RNA-sequencing (RNA-seq) methods only measure RNA species at a steady state and do not provide RNA dynamic information. Here, we develop addition-elimination mechanism-activated nucleotide transition sequencing (AENT-seq) for transcriptome-wide profiling of RNA dynamics. In AENT-seq, nascent transcripts are metabolically labeled with 4-thiouridine (4sU). The total RNA is treated with N2H4·H2O under aqueous conditions. N2H4·H2O is demonstrated to convert 4sU to 4-hydrazino cytosine (C*) based on an addition-elimination chemistry. C* is regarded as cytosine (C) during the DNA extension process. This 4sU-to-C transition marks nascent transcripts, so it enables sequencing analysis of RNA dynamics. We apply our AENT-seq to investigate transcript dynamic information of several genes involved in cancer progression and metastasis. This method uses a simple chemical reaction in aqueous solutions and will be rapidly disseminated with extensive applications.
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