NAP-seq用于全长非封顶RNA测序.
Shurong Liu1, Junhong Huang1, Lianghu Qu2
1MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, Guangdong Provincial Key Laboratory of Pharmaceutical Functional Genes, Innovation Center for Evolutionary Synthetic Biology, School of Life Sciences, Sun Yat-sen University, Guangzhou, China.
Nature protocols
|September 17, 2025
概括
非封闭RNAs (napRNAs) 是丰富的,但很难研究. NAP-seq是一种测序全长napRNA的新方法,有助于发现新型调节RNA和RNA生物发生路径.
科学领域:
- 基因组学就是基因组学.
- 分子生物学分子生物学
- 生物化学 生物化学
背景情况:
- 哺乳动物基因组广泛转录非封闭RNA (napRNAs).
- napRNAs调节生物过程和RNA生物发生,但由于它们的异质性,很难识别.
- 现有的方法与napRNA的不同长度,修改和结构作斗争.
研究的目的:
- 介绍napRNA测序 (NAP-seq) 技术的原理和详细程序.
- 为了能够在各种细胞类型中全面识别全长的napRNA.
- 克服napRNA发现和表征方面的挑战.
主要方法:
- NAP-seq使用T4多核酸激酶预处理来标准化RNA终结.
- 尺寸选择和RNase H枯竭丰富了长,低丰富度的RNA.
- 具有随机条形码的自定义适配器,双平台测序 (牛津纳米孔和Illumina) 和优化的cDNA合成确保单核酸分辨率并最大限度地减少偏差.
主要成果:
- NAP-seq允许在单核酸分辨率下识别具有各种终端修饰的全长napRNA.
- 这种方法有效地丰富了低丰度,长RNAs.
- 整个工作流,从图书馆准备到分析,可以在8天内完成.
结论:
- NAP-seq为发现具有调节作用的新型非编码RNA提供了强大的方法.
- 这种技术有助于在各种生物环境中研究RNA生物发生.
- NAP-seq克服了之前在napRNA识别和表征方面的局限性.
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