Flexible Method for Targeted Transcript Depletion from RNA-Seq Libraries: Mouse, Rat, Drosophila and Arabidopsis

This poster describes a novel method, Insert Dependent Adapter Cleavage (InDA-C), for effective removal of specific transcripts from RNA-Seq libraries without impacting non-targeted transcripts. InDA-C employs specific and robust enzymatic steps to eliminate undesirable transcripts such as rRNA during library construction without perturbing the original total RNA population as with hybridization capture methods. The specificity of transcript depletion relies on InDA-C primers which can be designed to target virtually any class of unwanted transcripts from any species. The library construction workflow uses as little as 10 ng of input total RNA, produces a strand-specific library, and is highly adaptable for depletion of any unwanted transcript(s).  Here we report the unbiased removal of rRNA and chloroplast RNA from RNA-Seq libraries across a variety of important model organisms including Mouse, Rat, Drosophila and Arabidopsis. Use of InDA-C primers designed against cytoplasmic + mitochondrial rRNAs from these species resulted in the reduction of sequencing reads derived from these transcripts to 2.5 - 8.7% of the total mapped reads. In the case of Arabidopsis, the combined use of InDA-C primers targeting rRNA and chloroplast rRNA resulted in an increase of informative reads (non-rRNA and non-Chloroplast rRNA) from 8.5% in absence of InDA-C depletion to 57% following targeted depletion with InDA-C. Therefore, through the use of InDA-C technology a greater percentage of RNA-Seq sequencing reads can be directed towards desired coding and non-coding transcripts.