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© Artur Scherf
Scanning Electron Microscopy of Red Blood Cell infected by Plasmodium falciparum.
Publication : Cellular microbiology

First efficient CRISPR-Cas9-mediated genome editing in Leishmania parasites

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in Cellular microbiology - 19 Jun 2015

Sollelis L, Ghorbal M, MacPherson CR, Martins RM, Kuk N, Crobu L, Bastien P, Scherf A, Lopez-Rubio JJ, Sterkers Y

Link to Pubmed [PMID] – 25939677

Cell. Microbiol. 2015 Oct;17(10):1405-12

Protozoan pathogens that cause leishmaniasis in humans are relatively refractory to genetic manipulation. In this work, we implemented the CRISPR-Cas9 system in Leishmania parasites and demonstrated its efficient use for genome editing. The Cas9 endonuclease was expressed under the control of the Dihydrofolate Reductase-Thymidylate Synthase (DHFR-TS) promoter and the single guide RNA was produced under the control of the U6snRNA promoter and terminator. As a proof of concept, we chose to knockout a tandemly repeated gene family, the paraflagellar rod-2 locus. We were able to obtain null mutants in a single round of transfection. In addition, we confirmed the absence of off-target editions by whole genome sequencing of two independent clones. Our work demonstrates that CRISPR-Cas9-mediated gene knockout represents a major improvement in comparison with existing methods. Beyond gene knockout, this genome editing tool opens avenues for a multitude of functional studies to speed up research on leishmaniasis.

http://www.ncbi.nlm.nih.gov/pubmed/25939677