ScriptCap™ m⁷G Capping System

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EPICENTRE's ScriptCap™ m⁷G Capping System is designed to build the Cap 0 structure found on the 5' end of most eukaryotic mRNA molecules. Based upon the tri-functional Vaccinia Virus capping enzyme (VCE), this system includes all of the components necessary to convert RNA containing a 5' triphosphate to Cap 0 RNA.

Unlike co-transcriptional capping systems which have theoretical capping yields of less than 80% capped message, the ScriptCap™ Capping System RNA exhibits almost 100% capped RNA. This RNA will also have all of the 5' caps in the proper orientation, eliminating another problem commonly associated with co-transcriptional systems.

The capping of in vitro transcribed RNA improves the stability and in vivo translation efficiency of transfected mRNA in most eukaryotic cells. Capped mRNA is also more efficiently translated in some in vitro translation systems.

Advantages of the ScriptCap include:

• 100% capped RNA. The theoretical maximum for most co-transcriptional capping systems is < 80%.
• Cap 1 structure. When used in conjunction with ScriptCap™ 2'-O-Methyltransferase, the natural Cap 1 structure can be built. This is the only commercially available system capable of building this translation-improving structure.
• Proper cap orientation. Unlike many traditional co-transcriptional cap analog systems, all ScriptCap™ built caps are in the proper orientation, improving in vivo translation efficiency.
• High yields. Since the RNA transcripts are produced using standard IVT systems, much higher yields of RNA are possible than in co-transcriptional systems.
• Cost. Better mRNA at a lower cost than co-transcriptional capping kits. No need to purchase expensive cap analogs.

Figure 1: Denaturing polyacrylamide gelstained with ethidium bromide. Lane 2 contains an uncapped, in vitro transcribed 51 base transcript. Lane 3 shows the addition of a single base due to VCE, which is consequently removed in Lane 4 by tobacco acid pyrophosphatase (TAP). Lane 5 shows that the capped transcripts are efficiently tailed using the A-Plus™ Poly(A) Tailing Kit.

Figure 2: (A) Denaturing Polyacrylamide gel and (B) autoradiograph of a ScriptCap Capping system reaction. Lane 2 shows the uncapped transcript without Vaccinia capping enzyme (VCE). Lane 3 includes the VCE and 14C SAM. VCE has clearly transferred the guanine base (Lane 3A) and the 14C containing methyl group (Lane 3B).