Fig. 1

Activation of the HSE promoter in medaka embryos and in cell culture. The gfp:HSE:luc DNA construct was co-injected with meganuclease into one-cell stage medaka embryos (A and B) or transfected in mouse Cop8 cells (C, C2, D, and D2). The injected embryos were divided into a control group and a test group. Embryos of the untreated control group (A) were gfp-negative. Embryos of the test group were treated at 39°C for 2 h, resulting in strong gfp expression (B). Typical embryos (stage 24) are shown for both groups. After transfection into mouse Cop8 cells, control plates remained gfp-negative (C). Treatment at 44°C for 2 h induced a strong gfp response in the transfected cells (D). (C2 and D2) are the corresponding brightfield views for C and D, respectively. (A, B, C, and D) are fluorescent images, background light was added for image (A) to visualize the otherwise gfp-negative embryo. A schematic presentation of the HSE promoter is depicted in (E). The artificial promoter contains eight multimerized heat shock elements flanked by two minimal promoters in opposed orientation (black arrowhead). Gfp on one side and the gene of interest (luciferase or Fgf8) at the other side are expressed from the bicistronic promoter. The vector is flanked by I-SceI meganuclease sites (arrows). Abbreviations: od, oil droplet; pA, SV40 polyadenylation signal; HSE, heat shock element; g.o.i., gene of interest.