Cre-Controlled CRISPR (3C) mutagenesis allows gene inactivation in Cre-dependent manner.a Scheme of the 3C rationale. A Cre effector construct controls the expression of a floxed Stop cassette upstream of the sequence encoding a fusion protein of Cas9 and GFP. In addition, a U6a promoter drives the constitutive expression of a gRNA targeting a gene of interest (GOI). Following exogenous or transgenic Cre supply, site-specific recombination results in the expression of Cas9-GFP. Combined with the gRNA a functional CRISPR complex is formed and mutates the target site within the gene of interest. b Scheme of the 3C gene inactivation construct targeting tyrosinase (tyr). The temperature-inducible hsp70l promotor drives expression of a floxed DsRed cassette. c Identification of transgenic animals expressing DsRed at 50 hpf after a heat treatment at 24 hpf. Example shown is a representative of a total of >100 heat-treated clutches from four independent 3C tyr transgenic insertions. Scale bar: 1000 µm.
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