Analysis and design of RNA sequencing experiments for identifying isoform regulation
Author(s)
Katz, Yarden; Airoldi, Edoardo M.; Wang, Eric T; Burge, Christopher B
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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.
Date issued
2010-11Department
Harvard University--MIT Division of Health Sciences and Technology; Massachusetts Institute of Technology. Department of Biological Engineering; Massachusetts Institute of Technology. Department of Biology; Massachusetts Institute of Technology. Department of Brain and Cognitive SciencesJournal
Nature Methods
Publisher
Nature Publishing Group
Citation
Katz, Yarden, Eric T Wang, Edoardo M Airoldi, and Christopher B Burge. “Analysis and design of RNA sequencing experiments for identifying isoform regulation.” Nature Methods 7, no. 12 (November 7, 2010): 1009-1015.
Version: Author's final manuscript
ISSN
1548-7091
1548-7105