Fig. 4. The easy-to-screen white platform for the yellow mutagenesis.

(A) Schematic representation of the generation of HDR-mediated yellow mutagenesis using the easy-to-screen white platform. The cartoon scissors indicate the TALENs and their cutting site at the yellow locus. The pair of TALEN binding sites is marked by the boxes. The empty triangles are two FRT sites located in the 5′ regulatory region and the intron of the white gene, and are oriented in the same direction. The two red-filled boxes indicate two genomic parts of the white gene separated by the second FRT site. Arrows with the names of L^F, L^R, R^F, R^R indicate the primers used for PCRs to get the L (left) and R (right) fragments as indicated in panel B. (B) Molecular identification of the white knock-in at the yellow locus, resulting in a simultaneous mutation in yellow. The genomic DNA of heterozygous F₁ line, 61-1, was used for showing the positive PCR results. L^F and L^R were used as primers to get the L fragment. R^F and R^R were used as primers to get the R fragment. (C–E) Removal of the white⁺ marker carried in the yellow mutants. (C) An eye of Lig4¹⁶⁹ flies. (D) An eye of the yellow mutant line, 61-1, which carried the white knock-in. (E) A mosaic white eye phenotype induced by heat-shock on the offspring of line 61-1 that was crossed with hs-Flp. See Fig. 1 legend for the common elements or labels not explained here.