FLP RECOMBINASE IN TRANSGENIC PLANTS - CONSTITUTIVE ACTIVITY IN STABLY TRANSFORMED TOBACCO AND GENERATION OF MARKED CELL CLONES IN ARABIDOPSIS

FLP site-specific recombinase was expressed in stably transformed toba cco and Arabidopsis. FLP-expressing tobacco lines were crossed with ot her transformed tobacco lines that contained a stably integrated FLP r ecognition target construct(s). The target construct consisted of two directly-oriented FLP recognition targets (FRTs), flanking a hygromyci n resistance cassette located between a GUS coding region and an upstr eam 35S CaMV promoter. Excision of the hygromycin resistance cassette by FLP-mediated recombination between FRTs brings the GUS coding regio n under the transcriptional control of the CaMV 35S promoter. In the a bsence of FLP-mediated recombination, the GUS gene is transcriptionall y silent. GUS activity was observed in the progeny of all crosses made between FLP recombinase-expressing and target-containing tobacco line s, but not in the selfs of parents. The predicted recombination produc t remaining after excision was confirmed by PCR and Southern analysis. In Arabidopsis, inducible expression of FLP recombinase was achieved from the soybean Gmhsp 17.6L heat-shock promoter. Heat-shock induction of FLP expression in plants containing the target construct led to ac tivation of constitutive GUS expression in a subset of cells, whose pr ogeny, therefore, were GUS-positive. A variety of clonal sectors were produced in plants derived from seed that was heat-shocked during germ ination. The ability to control the timing of GUS activation was demon strated by heat-shock of unopened flower heads which produced large se ctors. It was concluded that heat-shock-induced expression of FLP reco mbinase provides a readily controllable method for generating marked c lonal sectors in Arabidopsis, the size and distribution of which refle cts the timing of applied heat-shock.