Sequence-selective purification of biological RNAs using DNA nanoswitches

Abstract

SummaryNucleic acid purification is a critical aspect of biomedical research and a multibillion-dollar industry. Here we establish sequence-selective RNA capture, release, and isolation using conformationally responsive DNA nanoswitches. We validate purification of specific RNAs ranging in size from 22 to 401 nt with up to 75% recovery and 99.98% purity in a benchtop process with minimal expense and equipment. Our method compared favorably with bead-based extraction of an endogenous microRNA from cellular total RNA, and can be programmed for multiplexed purification of multiple individual RNA targets from one sample. Coupling our approach with downstream LC/MS, we analyzed RNA modifications in 5.8S ribosomal RNA, and found 2′-O-methylguanosine, 2′-O-methyluridine, and pseudouridine in a ratio of ∼1:7:22. The simplicity, low cost, and low sample requirements of our method make it suitable for easy adoption, and the versatility of the approach provides opportunities to expand the strategy to other biomolecules.