Transformation of chrysanthemum with mutated ethylene receptor genes: mDG-ERS1 transgenes conferring reduced ethylene sensitivity and characterization of the transformants

We generated mutated ethylene receptor genes (mDG-ERS1s) from the chrysanthemum ethylene receptor (DG-ERS1) cDNA by introducing one-nucleotide substitutions corresponding to those present in Arabidopsis etr1-1, etr1-2, etr1-3, and etr1-4 and tomato Nr. The promoter of a tobacco elongation factor 1α (EF1α) gene was fused to DG-ERS1 cDNA or one of the mDG-ERS1s. The resulting constructs were named EF1α∷mDG-ERS1(etr1-1), -ERS1(etr1-2) and so on, and introduced into chrysanthemum cv. Sei-Marine. We obtained putative transformants resistant to an antibiotic paromomycin with a yield of 2.4-6.2% depending on the construct. The mDG-ERS1(etr1-4) construct tended to be more effective in conferring reduced ethylene sensitivity in chrysanthemum than the others. PCR analysis gave amplification corresponding to a partial sequence of EF1α∷mDG-ERS1 transgenes. Southern blot analysis showed that, in the mDG-ERS1(etr1-4) transformant, not only the lines with reduced sensitivity to ethylene but also those sensitive to ethylene harbored the mDG-ERS1(etr1-4) transgene. The present results showed the usefulness of mutated ethylene receptor genes mDG-ERS1s for generation of transgenic chrysanthemums with reduced ethylene sensitivity.