Figure 5.

In vitro assay of methylation of synthetic miR-21-5p by HEMT1. The methylation assay is carried out in 20 μl reaction buffer containing 10 μM synthetic miR-21-5p and 5μM HENMT1 recombinant proteins. After incubating at 37°C for 40 min, RNA was recovered and purified by ethanol precipitation. (A–E) Methylation of synthetic miR-21-5p at 3′-end by purified HENMT1 assessed by EPIgeneous™ methyltransferase assay (A), Northern blot (B), qRT-PCR (C), mass spectrometry (D) and LC–MS/MS (E), respectively.