The Use of the Emu Promoter With Antibiotic and Herbicide Resistance Genes for the Selection of Transgenic Wheat Callus and Rice Plants

Abstract

Coding sequences of selectable marker genes were introduced into a plasmid containing the Emu promoter sequence which drives high levels of expression in cereal cells. The selectable markers were those coding for the enzymes neomycin phosphotransferase (NPT*) for kanamycin resistance, hygromycin phosphotransferase (HPT) for hygromycin resistance, phosphinothricin N-acetyltransferase (PAT) for resistance to the herbicidal compound phosphinothricin, and a mutant acetolactate synthase (ALS) for resistance to sulfonylurea herbicides. Plasmid DNAs were introduced into wheat and rice protoplasts by electroporation. Wheat protoplast-derived callus lines producing NPT and PAT were selected with kanamycin and phosphinothricin respectively. Wheat callus transformed with the gene coding for the mutant ALS was selected with sulfometuron methyl following the bombardment of suspension cultures with DNA-coated microparticles. With rice, transformed plants were regenerated from protoplast-derived cultures selected with kanamycin and hygromycin. Integration of the introduced DNA in transformed rice plants was confirmed by gel blot hybridisation, and significant levels of the NPT enzyme were measured by ELISA in leaves of the primary transformants as well as in leaves, seeds and roots of progeny plants. The results confirm that efficient selection of transformed cereal cultures and plants can be achieved by using selectable genes linked to the Emu promoter.