3´-O-Me-m7G(5')ppp(5')G RNA Cap Structure Analog
- Also known as Anti-Reverse Cap Analog (ARCA)
- Produces 100% translatable capped transcripts
- Can be used for co-transcriptional capping with T7 (NEB #M0251), SP6 (NEB #M0207) and T3 RNA Polymerases
- Can be used for synthesis of m7G capped RNA for in vitro splicing assays
- Can be used for synthesis of m7G capped RNA for transfection or microinjection
- Supplied as a lyophilized sodium salt
Avoiding RNase Contamination
Blocking of the 3´ -hydroxyl of m7G with 3´-0-Me assures that the capped transcripts are homogeneous. The 3´ - hydroxyl of the non-methylated G is the only 3´ - hydroxyl available for initiation (1).
The 5´terminal m7G cap present on most eukaryotic mRNAs promotes translation in vitro at the initiation level (2,3,4). For most RNAs, elimination of the cap structure causes a loss of stability, especially against exonuclease degradation (5), and a decrease in the formation of the initiation complex of mRNAs for protein synthesis (5,6). Certain prokaryotic mRNAs containing a 5´ terminal cap structure are translated as efficiently as or more efficiently than eukaryotic mRNAs in a eukaryotic cell-free protein synthesizing system (6). Also a cap requirement has been observed for splicing eukaryotic substrate RNAs (7).
A method using E. coli RNA Polymerase primed with m7G(5´)ppp(5´)G or m7G(5´)ppp(5´)A for an efficient in vitro synthesis of capped RNAs has been developed by Contreas (8). Larger amounts of capped RNAs are produced by transcription systems using SP6 RNA Polymerase primed with m7G(5´)ppp(5´)G (7).
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