The biochemistry of RNA-Seq library preparation results in cDNA fragments that are not uniformly distributed within the transcripts they represent. This non-uniformity must be accounted for when estimating expression levels, and we show how to perform the needed corrections using a likelihood based approach. We find improvements in expression estimates as measured by correlation with independently performed qRT-PCR and show that correction of bias leads to improved replicability of results across libraries and sequencing technologies.
Roberts A, Trapnell C, Donaghey J, Rinn JL, Pachter L. (2011) Improving RNA-Seq expression estimates by correcting for fragment bias. Genome Biol 12(3), R22. [Epub ahead of print]. [abstract]