Göttgens B , Nastos A , Kinston S , Piltz S , Delabesse EC , Stanley M , Sanchez MJ , Ciau-Uitz A , Patient R , Green AR .

Fig. 4. The +19 core enhancer is active in transgenic frogs and murine haematopoietic progenitor cells and critically depends on regions 1, 2 and 3. (A) An example of the variable ectopic expression patterns seen in a minority of transgenic stage 32 embryos with the SV/GFP construct (ectopic expression was never seen in the DLP region). (B) SV/GFP/H2.5 transgenic embryo showing reporter gene expression in the DLP region (arrow). (C–E) Stage 27 SV/GFP/H2.5 transgenic embryo stained for GFP (BCIP, turquoise) and endogenous SCL (BM Purple). d/e indicates the plane of the section shown in (D) and (E) which demonstrate that the purple staining (endogenous SCL) is contained within the area of transgene expression (turquoise) in the DLP (arrow). (F) Summary of transgenic frog experiments using wild-type and mutant versions of the SH0.6 core enhancer. DLP, embryos showing DLP staining with or without ectopic expression; ectopic, embryos showing ectopic expression only. The number in each bar represents the number of GFP-expressing embryos for each category. (G) The SV/luc/SH0.6 construct was active in 416B but not BW5147 cells in stable transfection assays. (H) Stable transfection assays in 416B cells demonstrate that the SH0.6 enhancer activity critically depends on the 227 bp 3′ fragment deleted in the SA0.4 construct, and that the three conserved regions mutated in the mut1, mut2 and mut3 constructs are each critical for enhancer function.

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