FIG. 3.

Localization of the MMTV pro-B-cell enhancer within the Mtv-1 LTR by deletion analysis in Ba/F3 cells. The enhancer test plasmid pGL2-Promoter with the Mtv-1 LTR fragment from positions −1162 to −613 cloned into the unique XhoI site 5′ of the promoter-reporter gene cassette in either the plus (pGL2-E5⁺, filled squares) or minus (pGL2-E5⁻, open triangles) orientation and mutant plasmids with truncations to positions −1025, −972, −840, −750, and −737 for pGL2-E5⁺ and to positions −761, −838, −893, −969, −1096, and −1154 for pGL2-E5⁻ were linearized with BamHI and transfected into Ba/F3 cells by electroporation. For each plasmid, a total of four transfections with two different plasmid preparations was done. After 30 h, cells were harvested and lysed and luciferase activity was determined as described in the legend to Fig. 2. Results are expressed as relative light units and represent the arithmetic means ± standard deviations from at least three parallel experiments with two different DNA preparations. Shown below the graph are a diagram of the MMTV (Mtv-1) enhancer fragment from positions −1162 to −613 as described previously (20) and a representation of the MMTV LTR with the functional regions (U3, R, and U5), transcription start site (+1) (labeled by a bent arrow), and restriction sites indicated. A hatched box indicates the region required for full activity.