Isoxaflutole treatment leads to reversible tissue bleaching and allows for more effective detection of GFP in transgenic soybean tissues

The effects of the herbicide isoxaflutole (IFT) on tissue growth and the detection of green fluorescent protein (GFP) in transgenic embryogenic soybean tissues were evaluated using image analysis. The inclusion of this "bleaching" herbicide at 3 or 10 mg l(-1) in a standard soybean embryo proliferation medium resulted in a change in tissue color from green to non-pigmented over the course of a 4-wk experiment. Although the loss in pigmentation was observed in transgenic and non-transformed control tissues, tissue growth remained unaffected. GFP expression in three different transgenic soybean clones, representing low to moderate GFP expression levels, was easily detected and quantified using image analysis following culture of the tissues on an IFT-containing medium. Quantification of GFP in tissues from the same clones cultured in the absence of IFT, however, was difficult using image analysis. After transfer of transgenic embryogenic tissue from a medium containing IFT to a medium without IFT, the growth of pigment-containing tissue resumed. The bleaching effects from this herbicide appear to be reversible and make IFT and possibly other bleaching herbicides useful in the analysis of GFP expression in tissues, where interference from chlorophyll is problematic.