Template production for mRNA in vitro transcription by PCR v1
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.
Authors
- Christian CR Renicke (ORCID: https://orcid.org/0000-0003-3226-9840)
Publication Details
- Published
- 2025-06-20
- DOI
- https://doi.org/10.17504/protocols.io.bp2l6yo4zvqe/v1
- Primary Topic
- Molecular Biology Techniques and Applications
- Type
- article
- Field-Weighted Citation Impact
- 0.00