Scheme of the transgenic heat-inducible VEGF zebrafish. (A) The pKTol2H70-mC-hVEGF-gcG transgene composed of a heat-inducible HSP70 promoter driving a floxed mCherry gene and hVEGF and a γ-crystallin promoter driving EGFP. mTol2 ITR indicates mini-Tol2 plasmid inverted terminal repeat; BGH(A), bovine growth hormone polyadenylation signal; SV40(A), simian virus 40 polyadenylation signal; rβG(A), rabbit β-globin polyadenylation signal. (B) The HSP70 promoter drives transcription of the mCherry gene, producing a red fluorescent protein in transgenic zebrafish. The lens-specific γ-crystallin promoter drives EGFP in the eyes. mC indicates mCherry; HS, heat shock. (C) Microinjection of cre recombinase mRNA into single-cell embryos results in the excision of the floxed mCherry gene, and subsequent heat-shock induction of the HSP70 promoter produces hVEGF. (D-E) Noninjected and cre recombinase–injected transgenic zebrafish were heat shocked at 37°C at 3 dpf to monitor the temporal expression of mCherry (D) and hVEGF (E) transcripts after induction of HSP70.