DNA-independent amplification of diverse RNA sequences by bacteriophage transcription polymerases

RNA replication and amplification have broad applications across biomedicine, but current methods are limited by a reliance on inefficient, multi-step protocols. Researchers at Stanford have developed a method for rapid, isothermal replication of RNA using a DNA-dependent RNA polymerase, T7 RNAP, producing large amounts of RNA from a single reaction without a DNA intermediate.

The inventors have characterized template requirements for RNA replication by T7 RNAP, and found that a diversity of RNAs with particular structures can be rapidly amplified. Under a different set of reaction conditions, a DNA 'seed' can also be used to evolve novel replicating RNA templates.

The inventors have further demonstrated the capability to conduct these reactions using a microfluidic setup, providing a means to more uniformly amplify a diverse pool of RNAs while mitigating amplification of contaminating templates.