Site-specific analysis of ribosomal 2′O-methylation by quantitative reverse transcription PCR under low deoxynucleotide triphosphate concentrations

Abstract

Ribose 2′O-methylation (Nm, ribomethylation) is the most abundant RNA modification present in rRNA. It has been shown that alterations in ribosomal 2′O-methylation at individual Nm sites likely reflect regulated cellular processes. Although several analytical approaches for Nm detection and profiling have been developed, a simple and affordable method for the screening and measurement of individual Nm sites in large numbers of tissue samples is required to examine their potential for clinical translation. Here, we describe a new quantitative reverse transcription PCR-based method that can sensitively assess ribomethylation levels at specific rRNA sites at single-nucleotide resolution in low input amounts of total RNA.

Keywords

• 2′O-methylation
• C/D-box snoRNA
• epitranscriptomics
• quantitative real-time PCR
• ribosomal RNA