Analysis and design of RNA sequencing experiments for identifying isoform regulation.
Analysis and design of RNA sequencing experiments for identifying isoform regulation.
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DOI：
10.1038/nmeth.1528
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发表时间：
2010-12
期刊：
影响因子：
48
通讯作者：
Burge, Christopher B.
中科院分区：
文献类型：
--
作者：
Katz, Yarden;Wang, Eric T.;Airoldi, Edoardo M.;Burge, Christopher B.
通过选择性剪接，大多数人类基因表达多个同种型，这些同种型通常在功能上不同。为了从cDNA片段的高通量测序（RNA-seq）中推断亚型调控，我们开发了混合亚型（MISO）模型，这是一种估计可变剪接外显子和亚型表达的统计模型，并评估这些估计的置信度。在配对末端RNA-seq中纳入mRNA片段长度分布大大改善了对选择性剪接水平的估计。MISO还检测差异调节的外显子或同种型。MISO的应用涉及RNA剪接因子hnRNP H1调节选择性切割和聚腺苷酸化，这一作用得到了人类细胞中UV交联免疫沉淀测序（CLIP-seq）分析的支持。我们的研究结果为RNA-seq分析提供了一个概率框架，为前mRNA加工提供了功能性见解，并为基因和亚型表达研究的RNA-seq实验的最佳设计提供了指导。
Through alternative splicing, most human genes express multiple isoforms that often differ in function. To infer isoform regulation from high-throughput sequencing of cDNA fragments (RNA-seq), we developed the mixture-of-isoforms (MISO) model, a statistical model that estimates expression of alternatively spliced exons and isoforms and assesses confidence in these estimates. Incorporation of mRNA fragment length distribution in paired-end RNA-seq greatly improved estimation of alternative-splicing levels. MISO also detects differentially regulated exons or isoforms. Application of MISO implicated the RNA splicing factor hnRNP H1 in the regulation of alternative cleavage and polyadenylation, a role that was supported by UV cross-linking–immunoprecipitation sequencing (CLIP-seq) analysis in human cells. Our results provide a probabilistic framework for RNA-seq analysis, give functional insights into pre-mRNA processing and yield guidelines for the optimal design of RNA-seq experiments for studies of gene and isoform expression.
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影响因子：
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