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Harnessing tRNA for Processing Ability and Promoter Activity.

Knapp DJHF, Fulga TA

Radcliffe Department of Medicine, Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, UK. david.knapp@umontreal.ca.

Transfer RNA (tRNA) and their associated production and processing machinery can be coopted as a versatile tool for the production of guide RNAs (gRNAs) for Cas9-based genome engineering. Using different tRNA variants enables the production of gRNAs at a variety of steady state levels. Furthermore, engineered tRNAs can be used to process gRNAs from Pol-II transcripts, thus enabling spatial/temporal control of gRNA expression. Here we describe the design, cloning, and testing of tRNA scaffolds for both Pol-III-driven expression of different levels of gRNAs, and for processing gRNAs from Pol-II transcripts.

Methods Mol Biol 2021;2162:89-114.

Pubmed ID: 32926380

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