Bisulfite-sequencing analysis of previously reported m5C sites in MLV RNA. (A) Schematic representation of MLV genome structure. Amplicon sequences harboring four previously reported (Eckwahl et al. 2020) m5C candidate sites (marked in red) toward the 5′ end (left) and the 3′ end (right), respectively, are shown (numbers corresponding to NC_001362.1 sequence). (B) Bisulfite-treated RNA from virus-infected Mus dunni cells was reverse transcribed, and the regions indicated in (A) were amplified by PCR followed by subcloning and Sanger sequencing of individual clones (n = 35 each). No unconverted Cs were detected at the indicated positions in viral RNA. Methylated positions C38, C47, and C48 in mouse tRNAAsp are shown as positive control (n = 10); results from the spike-in ERCC136 RNA (nt 81–323) are shown as negative control (n = 10).
