While post-transcriptional modifications are a characteristic feature of noncoding RNAs, the biological function of these modifications is unknown. Cytosine-5 methylation has been detected in abundant RNA molecules including ribosomal RNAs and transfer RNAs, but the methylation status of cytosines in other noncoding RNAs is not known. To further investigate these modifications, Matthias Schaefer and colleagues from the German Cancer Research Center have developed a protocol for Detection of Cytosine Methylation in RNA Using Bisulfite Sequencing. The method, featured in the October issue of Cold Spring Harbor Protocols, uses a bisulfite treatment of RNA to chemically deaminate nonmethylated cytosines to uracil, leaving methylated cytosines unaffected. Subsequent cDNA synthesis and PCR amplification offers researchers material for high-throughput sequencing analysis of the methylation patterns in any RNA molecule, including noncoding RNAs and low-abundance RNAs. As one of October’s featured articles, the protocol is freely available to subscribers and nonsubscribers alike.