RNA-Seq of human whole blood: Evaluation of globin RNA depletion on Ribo-Zero library method

Peripheral blood is a highly accessible biofluid providing a rich source of information about human physiology and health status. However, for studies of the blood transcriptome with RNA sequencing (RNA-Seq) techniques, high levels of hemoglobin mRNAs (hgRNA) present in blood can occupy valuable sequencing space, impacting detection and quantification of non-hgRNAs. In this study, we evaluated two methods for preparing ribosomal RNA (rRNA)-depleted sequencing libraries for RNA-Seq of whole blood, one of which is also designed to deplete hgRNAs. Two experiments were performed: one evaluating library performance across 6 human blood samples and the other examining library reproducibility and performance in a two subject subset. We find that addition of hgRNA depletion to the rRNA-depletion protocol for library preparation from blood RNA effectively reduces highly abundant hgRNA reads in RNA-Seq data but does not result in an increase in detection of differentially expressed genes (DEG).