EFFICIENT RIBOSOMAL RNA DEPLETION FROM DROSOPHILA TOTAL RNA FOR NEXT-GENERATION SEQUENCING APPLICATIONS

We developed a cost-effective enzyme-based rRNA-depletion method tailored for Drosophila melanogaster, addressing the limitations of existing commercial kits and the lack of peer-reviewed alternatives. Our method employs single-stranded DNA probes complementary to Drosophila rRNA, forming DNA-RNA hybrids. These hybrids are then degraded using RNAse H enzyme, effectively removing rRNA and enriching all non-ribosomal RNAs, including mRNA, lncRNA and small RNA. When compared to a commercial rRNA Fly Kit, our approach demonstrated superior rRNA removal efficiency and mapping percentage, confirming its effectiveness. Additionally, our method successfully enriched the non-coding transcriptome, making it a valuable tool for studying ncRNA in Drosophila. The probe sequences and rRNA-depletion protocol is made freely available, offering researchers a reliable alternative for rRNA-depletion experiments. RNA-seq profiling of Drosophila adult brains processed by PolyA enrichment and rRNA depletion