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Modeling enzyme processivity reveals that RNA-Seq libraries are biased in characteristic and correctable ways
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Archer, Nathan, Walsh, Mark, Shahrezaie, Vahid and Hebenstreit, Daniel (2016) Modeling enzyme processivity reveals that RNA-Seq libraries are biased in characteristic and correctable ways. Cell Systems, 3 (5). pp. 467-479. doi:10.1016/j.cels.2016.10.012 ISSN 2405-4712.
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Official URL: http://dx.doi.org/10.1016/j.cels.2016.10.012
Abstract
Experimental procedures for preparing RNA-seq and single-cell (sc) RNA-seq libraries are based on assumptions regarding their underlying enzymatic reactions. Here, we show that the fairness of these assumptions varies within libraries: coverage by sequencing reads along and between transcripts exhibits characteristic, protocol-dependent biases. To understand the mechanistic basis of this bias, we present an integrated modeling framework that infers the relationship between enzyme reactions during library preparation and the characteristic coverage patterns observed for different protocols. Analysis of new and existing (sc)RNA-seq data from six different library preparation protocols reveals that polymerase processivity is the mechanistic origin of coverage biases. We apply our framework to demonstrate that lowering incubation temperature increases processivity, yield, and (sc)RNA-seq sensitivity in all protocols. We also provide correction factors based on our model for increasing accuracy of transcript quantification in existing samples prepared at standard temperatures. In total, our findings improve our ability to accurately reflect in vivo transcript abundances in (sc)RNA-seq libraries.
Item Type: | Journal Article | ||||||||||
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Divisions: | Faculty of Science, Engineering and Medicine > Science > Life Sciences (2010- ) | ||||||||||
Journal or Publication Title: | Cell Systems | ||||||||||
Publisher: | Cell Press | ||||||||||
ISSN: | 2405-4712 | ||||||||||
Official Date: | 23 November 2016 | ||||||||||
Dates: |
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Volume: | 3 | ||||||||||
Number: | 5 | ||||||||||
Page Range: | pp. 467-479 | ||||||||||
DOI: | 10.1016/j.cels.2016.10.012 | ||||||||||
Status: | Peer Reviewed | ||||||||||
Publication Status: | Published | ||||||||||
Access rights to Published version: | Open Access (Creative Commons) | ||||||||||
Description: | Accepted under title "RNA-seq data is shaped in characteristic ways by polymerase processiviities" |
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Date of first compliant deposit: | 8 September 2016 | ||||||||||
Date of first compliant Open Access: | 14 November 2016 | ||||||||||
Funder: | Royal Society (Great Britain), Biotechnology and Biological Sciences Research Council (Great Britain) (BBSRC) | ||||||||||
Grant number: | RG130085 (Royal Society ) BB/L006340/1 ; BB/M017982/1 (BBSRC) | ||||||||||
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