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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. © 2011 Roberts et al.; licensee BioMed Central Ltd.
Roberts, A., Trapnell, C., Donaghey, J., Rinn, J. L., & Pachter, L. (2011). Improving RNA-Seq expression estimates by correcting for fragment bias. Genome Biology, 12(3). https://doi.org/10.1186/gb-2011-12-3-r22