Poster: Emerging technologies

Abs # 916: Stable expression of yeast FLP site-specific recombinase in rice

Presenter:	Luo, Hong , hongluo@hybrigene.com
Authors	Hu, Qian  (B)   Viola, David  (A)   Zen, Peiyu  (A)   Kausch, Albert P (B) (A)  Chandlee, Joel M (A)   Luo, Hong  (B) (A)
Affiliations:	(A): University of Rhode Island (B): HybriGene, Inc.

We have evaluated the feasibility of using FLP/FRT site-specific recombination system in rice for genome engineering of crop species. The objective of our study was to produce transgenic rice lines stably expressing the yeast FLP recombinase and evaluate in vivo efficacy of FLP-mediated site-specific DNA recombination. We hypothesized that transgenic rice plants containing a recombination-reporter construct, in which the rice ubiquitin promoter and the reporter gusA coding region is separated by the hygromycin marker gene flanked by directly oriented FRT sites will not show GUS activity. When crossed with plants containing stably expressed FLP recombinase, FLP should excise the blocking fragment (hyg gene) thus bringing together the ubiquitin promoter and the downstream gusA gene, giving rise to GUS expression in the hybrid plant. Using Agrobacterium-mediated transformation, we obtained transgenic plants with the FLP-containing construct or the FRT-containing recombination-reporter construct. After cross-pollination between the FLP-expressing plants and FRT-containing plants, hybrid seeds were harvested and planted to produce F1 plants. While the majority of the hybrid progeny exhibited a uniform GUS expression, the progeny of selfed parental rice plants did not show detectable GUS activity. Molecular analysis further confirmed the FLP-mediated site-specific DNA excision. This observation demonstrates the efficient operation of FLP recombinase in catalyzing excisional DNA recombination in rice, indicating that the FLP/FRT recombination system functions in cereal crops.