Fig. 5.

TSG differential labeling of twin spots. (A) TSG is a Flp-FRT–based MR strategy, as in Fig. 2, except that the FRT site, located in an intron, is flanked with complementary GFP and RFP sequences to create TSG hybrid cassettes (Materials and Methods). (B) Heat shock-induced MR generates full-length but interrupted cds for GFP on one recombined chromosome and for RFP on the other. Splicing removes the FRT site along with the rest of the intron to reconstruct continuous full-length GFP or RFP cds. In a heterozygote, placing the wild-type allele distal to the 3′ RFP and the mutant allele distal to the 3′ GFP enables direct identification of both the red wild-type control and green mutant twins. (C) TSG provides a direct readout of toxicity. In control experiments in which both twin spots are homozygous wild type, statistical analysis showed equal numbers of cells present in the red and green twin spots, as expected and confirming that ectopic expression of GFP or RFP had no apparent toxic effects (compare with Fig. 6C). Adapted from ref. 39.