Codex^® HiCap RNA Polymerase

in vitro transcription for advancing RNA medicine

Codex® HiCap RNA Polymerase is a co-transcriptional capping RNA polymerase engineered to produce synthetic RNA to meet increasing demands for safety and cost-effectiveness. The HiCap enzyme generates fewer undesirable double-stranded RNA (dsRNA) byproducts than wild-type T7 (WT T7) RNA polymerase, reducing unwanted immunogenicity. In addition, the enzyme incorporates commercially available cap analogs more efficiently, reducing overall manufacturing costs.

dsRNA byproducts

Replicate IVTs were prepared using a ~2kb plasmid DNA template and either WT T7 or Codex®HiCap RNA Polymerase with Anti-Reverse Cap Analog (ARCA) or CleanCap AG. All IVTs were incubated for 2 hrs at 37°C . Both 2.5 and 25µg of the resulting raw mRNA were evaluated for presence of dsRNA byproducts by dot blot using J2 dsRNA specific primary antibody.

Capping Efficiency

WT T7 IVTs were prepared with 5mM ARCA, and 1mM GTP or 4mM CleanCap AG. Codex® HiCap RNA IVTs were prepared 4mM ARCA, and 5mM GTP or with 1.5 mM CleanCap AG. All IVTs were incubated for 2 hrs at 37°C .mRNA titers were assessed using the Invitrogen Quant-IT RNA BR Assay Kit. IVT samples were further purified using NEB Monach RNA Cleanup kit and evaluated by gel electrophoresis.

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