Transcriptomics analyses play pivotal roles in understanding the complex regulatory networks that govern cellular processes. The abundance of rRNAs, which account for 80-90% of total RNA in eukaryotes, limits the detection and investigation of other transcripts. While mRNAs and long non-coding RNAs have polyA(+) tails that are often used for positive selection, investigations of polyA(-) RNAs, such as circular RNAs, histone mRNAs, and small RNAs, typically require the removal of the abundant rRNAs for enrichment. Current approaches to deplete rRNAs for downstream molecular biology investigations are hampered by restrictive RNA input masses and high cost. To address these challenges, we developed rRNA Removal by RNase H (rRRR), a method to efficiently deplete rRNA from a wide range of human, mouse, and rat RNA inputs and qualities at a cost 10-20-fold cheaper than other approaches. We employed probe-based hybridization and enzymatic digestion to selectively target and remove rRNA molecules while preserving the integrity of non-rRNA transcripts. Comparison between rRRR to two commercially available approaches found that they had similar efficiencies at depleting rRNAs and comparable off-target effects. Our developed method provides researchers with a valuable tool for investigating gene expression and regulatory mechanisms across a wide range of biological systems at an affordable price that increases the accessibility for researchers to enter the field, ultimately advancing our understanding of cellular processes.

中文翻译：

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一种可扩展且经济高效的 rRNA 去除方法，可富集 RNA 用于分子生物学研究

转录组学分析在理解控制细胞过程的复杂调控网络方面发挥着关键作用。rRNA 的丰度占真核生物总 RNA 的 80-90%，限制了其他转录本的检测和研究。虽然 mRNA 和长非编码 RNA 具有常用于正选择的多聚 A(+) 尾部，但对多聚 A(-) RNA（例如环状 RNA、组蛋白 mRNA 和小 RNA）的研究通常需要去除丰富的 rRNA为了丰富。目前用于下游分子生物学研究的耗尽 rRNA 的方法受到限制性 RNA 输入质量和高成本的阻碍。为了应对这些挑战，我们开发了 RNase H 去除 rRNA (rRRR)，这是一种有效去除各种人类、小鼠和大鼠 RNA 输入和质量中的 rRNA 的方法，其成本比其他方法便宜 10-20 倍。我们采用基于探针的杂交和酶消化来选择性地靶向和去除 rRNA 分子，同时保留非 rRNA 转录物的完整性。rRRR 与两种市售方法的比较发现，它们在消耗 rRNA 方面具有相似的效率和类似的脱靶效应。我们开发的方法为研究人员提供了一个有价值的工具，以可承受的价格研究各种生物系统中的基因表达和调控机制，增加了研究人员进入该领域的机会，最终增进了我们对细胞过程的理解。