Optimization of CRISPR/Cas9-mediated gene disruption in Xenopus laevis using a phenotypic image analysis technique.

ABSTRACT: The CRISPR/Cas9 method has become popular for gene disruption experiments in Xenopus laevis. However, the experimental conditions that influence the efficiency of CRISPR/Cas9 remain unclear. To that end, we developed an image analysis technique for the semi-quantitative evaluation of the pigment phenotype resulting from the disruption of tyrosinase genes in X. laevis using a CRISPR/Cas9 approach, and then examined the effects of varying five experimental parameters (timing of the CRISPR reagent injection into developing embryos; amount of Cas9 mRNA in the injection reagent; total injection volume per embryo; number of injection sites per embryo; and the culture temperature of the injected embryos) on the gene disruption efficiency. The results of this systematic analysis suggest that the h

SUBMITTER: Tanouchi M 

PROVIDER: S-EPMC11520957 | biostudies-literature | 2022 May

REPOSITORIES: biostudies-literature