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bioRxiv · 10.1101/2021.06.21.449263

Gene editing in bacteria via SpCas9/sgRNA ribonucleoprotein complexes

Arroyo-Olarte, R. D.·Bravo-Rodriguez, R. N.·Morales-Rios, E.
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Abstract

Gene editing has been revolutionized by CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)-Cas technology in a variety of organisms. In bacteria, however, CRISPR-Cas still holds many caveats such as high toxicity of Cas9 and off-target editing effects. In this work we develop a system for the incorporation of Cas9/single guide RNA ribonucleoprotein complexes in bacteria and their successful application for gene editing via homologous recombination. Targeting of a green fluorescent protein (GFP) reporter allows for easy verification of gene-edition via conversion to blue-fluorescent protein (BFP), mediated by the well characterized 196T > C (Tyr66His) mutation.

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Arroyo-Olarte, R. D., Bravo-Rodriguez, R. N., Morales-Rios, E.. 2021-06-21. Gene editing in bacteria via SpCas9/sgRNA ribonucleoprotein complexes. https://doi.org/10.1101/2021.06.21.449263

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