RNase H-based analysis of synthetic mRNA 5′ cap incorporation

Enzymatic reactions involved in RNA 5′ capping. RNA transcripts generated by in vitro transcription (usually done using T7 RNA polymerase or its variants; reaction 0) contains a triphosphate group at the 5′ end (pppR-). The 5′ triphosphate group can be converted to the Cap-1 structure (m⁷GpppRm-) through four enzymatic reactions carried out by RNA capping enzymes such as vaccinia RNA capping enzyme (reactions 1 through 3) in conjunction with vaccinia cap 2′-O-methyltransferase (reaction 4).