Expression of mouse dihydrofolate reductase gene confers methotrexate resistance in transgenic petunia plants

Transgenic petunia plants containing an altered (Leu22----Arg22) mouse dihydrofolate reductase gene fused to the cauliflower mosiac virus 35S (CaMV 35S) promoter and nopaline synthase (nos) polyadenylation site were obtained by transforming petunia leaf disks with an Agrobacterium tumefaciens strain carrying the chimeric gene. Transformants were directly selected for and rooted on medium containing 1 microM methotrexate (MTX). The chimeric gene was present in the regenerated plants at one to three copies and produced the expected 950-nucleotide-long transcript based on Southern and Northern hybridization analyses, respectively. Leaf pieces from the regenerated transgenic plants were able to form callus when cultured on medium containing 1 microM MTX and were able to incorporate 32P into high-molecular-weight DNA in the presence of greater than 100 microM MTX, thus demonstrating that the chimeric mouse dhfr gene was fully functional and useful as a selectable marker in plant transformation experiments. To date, this is the first report of successful expression of a vertebrate gene in transformed plant cells.