A novel plasmid-based co-tethered transcription platform for high yield, high purity mRNA synthesis.

ABSTRACT: This work aims to improve RNA synthesis and manufacturing, exemplified by T7 RNA polymerase-driven in vitro transcription. We developed a novel, plasmid-compatible co-tethering strategy that functionally couples RNA polymerase to its promoter DNA immobilized on a solid matrix. As demonstrated recently, co-tethering enhances promoter binding, increases RNA yield, and suppresses RNA re-binding, especially under high-salt conditions, thereby reducing double-stranded RNA by-products. The system leverages asymmetric end-labeling of linearized plasmid DNA using a simple "Klenow fill-in" reaction with modified nucleotides, enabling stable attachment of DNA to both RNA polymerase and solid support (magnetic beads). The immobilized co-tethered polymerase-DNA complex supports efficient transcription

SUBMITTER: Mala P 

PROVIDER: S-EPMC12746099 | biostudies-literature | 2025 Nov

REPOSITORIES: biostudies-literature