Cre-mediated recombination in the green-to-red reporter line. In Tg (ef1a: loxP-GFP-loxP-dsRed2) embryos injected with 3 pg of Cre mRNA at one-cell stage (A), dsRed fluorescence is ubiquitously observed at 36 hpf. Embryos were injected with 3 pg of CreER^T2 mRNA, further incubated with embryo medium (B) or 1 μM cyclofen-OH (Ind) (C), and observed at 2 dpf. Variation of the recombination efficiency with the amount of injected CreER^T2 mRNA (in pg) and inducer concentration: 0 μM (blue triangle) or 3 μM (red square). The percentage of embryos exhibiting dsRed fluorescence was scored by epifluorescence microscopy at 2 dpf (D). The error bars represent statistical errors estimated as √p(1-p)/n. Scale bars: 100 μm. dpf, days postfertilization.

Photocontrol of CreER^T2 recombinase activity in Tg reporter line. Transgenic (ef1α:loxP-GFP-loxP-dsRed2) embryos were injected with 3 pg of CreER^T2 mRNA at the one-cell stage and further incubated with 3 μM of caged inducer (cInd). Photoactivation of cInd at the 3/6-somite stage resulted in dsRed expression observed here at 2 dpf. In epifluorescence microscopy, the non-UV illuminated embryos (A) do not express dsRed, whereas they do express dsRed in every cell upon global UV illumination (B), in few cells upon twophoton excitation in a single cell in the forming retina (D), or in a just formed somite (E). Scale bars: 100 μm. Kinetics of recombination was determined by quantitative PCR (C). Genomic DNA was prepared from 30 tg (ef1α:loxP-GFP-loxPdsRed2) embryos; Green: NI, noninjected embryos; blue: embryos injected at one-cell stage with 30 pg cre-ER^T2 mRNA, incubated with 3 μM of caged inducer (cInd), illuminated at 50% epiboly, and then fixed for DNA purification; red: embryos injected at one-cell stage with 30 pg cre mRNA and allowed to develop until 24 hpf. The average values (±standard error of the mean) from at least three independent experiments are presented; comparisons with t=0 were performed by a Student’s t-test. Values of p < 0.05 were considered to indicate statistical significance (***p < 0.001; **p < 0.01).