What type of study is this?

This is a Pre-Registered Trial study.

September 10, 2025

Template production for mRNA in vitro transcription by PCR v1

CRChristian CR RenickeJohns Hopkins University

Key Points

The protocol produces 1 to 5 µg of dsDNA ampicon for mRNA transcription, ensuring high yield.
Key steps involve using KOD polymerase for dsDNA synthesis and DpnI for removing plasmid DNA.
Instructions include practical purification of the DNA, making it suitable for high-quality, in vitro transcription.
Scalability allows researchers to adjust quantities easily for larger applications in mRNA production.

Abstract

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.

KI fragen

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Cite This Study

Christian CR Renicke (Fri,) studied this question.

synapsesocial.com/papers/68c189d99b7b07f3a061375d https://doi.org/https://doi.org/10.17504/protocols.io.bp2l6yo4zvqe/v1