Accurate quantification of 3 ′-terminal 2′-O-methylated small RNAs by utilizing oxidative deep sequencing and stem-loop RT-qPCR

In this study, we profiled 3′-terminal 2′-O-methyl plant miRNAs in the livers of rice-fed mice by oxidative deep sequencing and detected increasing amounts of plant miRNAs with prolonged oxidation treatment. We further compared the efficiency of stem- loop and poly(A)-tailed RT-qPCR in quantifying plant miRNAs in animal tissues and identified stem-loop RT-qPCR as the only suitable approach. Likewise, stem-loop RT-qPCR was superior to poly(A)-tailed RT-qPCR in quantifying 3′-terminal 2′-O-methyl piRNAs in human seminal plasma. In summary, this study established a standard procedure for quantifying the levels of 3′-terminal 2′-O-methyl miRNAs in plants and piRNAs. Accurate measurement of the 3′-terminal 2′-O-methylation of small RNAs has profound implications for understanding their pathophysiologic roles in biological systems.

Source: Frontiers of Medicine - April 1, 2022 Category: General Medicine Source Type: research