This protocol outlines the production of dsDNA templates for mRNA in vitro transcription, indicating efficient purification methods.
This a protocol for producing a linear, dsDNA amplicon with KOD hot-start high-fidelity DNA polymerase for subsequent use in in vitro transcription of mRNA using SP6 RNA polymerase. It includes removal of the circular, dam-methylated plasmid DNA template via DpnI digestion and instructions for purification and concentration of the final DNA. A single reaction should yield 1 to 5 µg DNA and the protocol can easily scaled up to produce larger amounts of high-quality, in vitro transcription-ready DNA.
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Christian CR Renicke (2025) studied this question.
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