SnapShot-Seq: a method for extracting genome-wide, in vivo mRNA dynamics from a single total RNA sample.
SnapShot-Seq: a method for extracting genome-wide, in vivo mRNA dynamics from a single total RNA sample.
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DOI：
10.1371/journal.pone.0089673
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发表时间：
2014
期刊：
影响因子：
3.7
通讯作者：
Springer M
中科院分区：
文献类型：
--
作者：
Gray JM;Harmin DA;Boswell SA;Cloonan N;Mullen TE;Ling JJ;Miller N;Kuersten S;Ma YC;McCarroll SA;Grimmond SM;Springer M
信使核糖核酸（mRNA）的合成、加工和降解涉及一系列复杂的分子步骤，人们对其尚未完全了解。由于这些步骤中的每一个RNA中间体都有有限的寿命，大量的机制和动态信息都编码在细胞总RNA中。在此我们报道了快照测序（SnapShot - Seq）技术的发展，这是一套计算方法，能够从细胞总RNA的单次RNA测序快照中确定前体mRNA合成、剪接、内含子降解以及mRNA降解的体内速率。快照测序能够检测剪接特定步骤速率的体内变化，并且它提供了前体mRNA合成速率的全基因组估计值，该估计值与通过标记新合成的RNA所获得的值相当。我们利用快照测序研究了后生动物基因表达水平固有双峰性的起源，我们的结果表明，这种双峰性部分是由于转录激活从高表达基因向其低表达邻近基因的溢出。快照测序极大地扩展了从标准RNA测序实验中可获得的信息。
mRNA synthesis, processing, and destruction involve a complex series of molecular steps that are incompletely understood. Because the RNA intermediates in each of these steps have finite lifetimes, extensive mechanistic and dynamical information is encoded in total cellular RNA. Here we report the development of SnapShot-Seq, a set of computational methods that allow the determination of in vivo rates of pre-mRNA synthesis, splicing, intron degradation, and mRNA decay from a single RNA-Seq snapshot of total cellular RNA. SnapShot-Seq can detect in vivo changes in the rates of specific steps of splicing, and it provides genome-wide estimates of pre-mRNA synthesis rates comparable to those obtained via labeling of newly synthesized RNA. We used SnapShot-Seq to investigate the origins of the intrinsic bimodality of metazoan gene expression levels, and our results suggest that this bimodality is partly due to spillover of transcriptional activation from highly expressed genes to their poorly expressed neighbors. SnapShot-Seq dramatically expands the information obtainable from a standard RNA-Seq experiment.
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