We have constructed a chimeric gene, consisting of a 2 kb DNA fragment from the shrunken promoter, the bacterial neomycin phosphotransferase II gene from Tn5 and the polyadenylation site of the octopin synthase gene derived from the t-DNA of Agrobacterium tumefaciens. The promoter fragment from Zea mays contains approximately 2 kb upstream sequences, the transcription start and 40 bp of exon 1 (W. Werr et al., EMBO J. 4:1373, 1985). This exon contains only untranslated sequences. The first ATG in the chimeric transcript is provided by the bacterial sequences and starts the open reading frame encoding the neomycin phosphotransferase II enzyme.
We have used this construction to transform Triticum monococcum protoplasts by polyethylene glycol treatment (H. Lorz et al., MGG 199:178, 1985). We found expression of this chimeric gene, that means NPTII activity, over a time period of at least 10 days. The maximum enzyme activity was determined 4 days after transformation. The enzyme activity was dependent on the presence of the maize promoter fragment within the construction. This result suggests that the maize promoter acts in the Triticum monococcum cells (W. Werr and H. Lorz, MGG, 1986, in press). For a detailed analysis of this maize promoter we have constructed a series of deletion mutants within the promoter fragment of the chimeric gene which will be tested for activity within this transient expression system.
Wolfgang Werr, Christoph Maas, Horst Lorz1 and Peter Starlinger
1Abt. Schell. Max-Planck-Institut
fur Zuchtungsforschung, D-5000 Koln
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