Instantaneous visual genotyping and facile site-specific transgenesis via CRISPR-Cas9 and phiC31 integrase.

ABSTRACT: Here, we introduce 'TICIT', targeted integration by CRISPR-Cas9 and integrase technologies, which utilizes the site-specific DNA recombinase - phiC31 integrase - to insert large DNA fragments into CRISPR-Cas9 target loci. This technique, which relies on first knocking in a 39-basepair phiC31 landing site via CRISPR-Cas9, enables researchers to repeatedly perform site-specific transgenesis at the exact genomic location with high precision and efficiency. We applied this approach to devise a method for the instantaneous determination of a zebrafish's genotype simply by examining its color. When a zebrafish mutant line must be propagated as heterozygotes due to homozygous lethality, employing this method allows facile identification of a population of homozygous mutant embryos even before the

SUBMITTER: Ma J 

PROVIDER: S-EPMC11391820 | biostudies-literature | 2024 Sep

REPOSITORIES: biostudies-literature