Figure 4. The NEXTFLEX® Rapid Directional RNA-Seq kit 2.0 delivers libraries containing low levels of rRNA contamination compared to the Competitor N kit. rRNA-depleted total RNA was isolated from Universal Human Reference RNA (Agilent #740000) using the NEXTFLEX® RiboNaut™ rRNA depletion kit (human, mouse, rat) and the Competitor N rRNA-depletion kit. Libraries were generated using the NEXTFLEX® Rapid Directional RNA-Seq kit 2.0 and the Competitor N’s library preparation kit. The resulting libraries were sequenced on the Illumina® MiSeq® sequencer using single-end mode (1×151 bp). The reads were trimmed using cutadapt and the percent of rRNA was determined by using bowtie2 to map reads to human rRNA. The NEXTFLEX® Rapid Directional RNA-Seq kit 2.0 demonstrated superior removal of 5S, 5.8S, 12S, 16S, 18S, and 28S rRNA species compared to the Competitor N kit.