Expression of a bifunctional green fluorescent protein (GFP) fusion marker under the control of three constitutive promoters and enhanced derivatives in transgenic grape (Vitis vinifera).
Activity of three constitutive promoters and enhanced derivatives in transgenic grape (Vitis vinifera L. cv. Thompson Seedless) was characterized using a bifunctional fusion marker containing the enhanced green fluorescent protein (EGFP) and neomycin phosphotransferase (NPTII) genes. Relative differences in transient GFP expression and stable transformation efficiencies were used to compare promoter activity. Expression patterns in transformed somatic embryos revealed that the ACT2 promoter from Arabidopsis thaliana, previously shown to be a strong constitutive promoter in A. thaliana and other species, failed to promote strong expression in grape. In contrast, a promoter isolated from cassava vein mosaic virus (CsVMV) supported high levels of transgene expression equivalent to those achieved using an enhanced double cauliflower mosaic virus (CaMV) 35S promoter. Duplication of the 5'-upstream enhancer region of the CsVMV promoter further enhanced its ability to increase transgene expression. However, the pattern of transgene expression driven by these two viral promoters was significantly different at the whole plant level. The enhanced double CaMV 35S promoter was highly active in most tissues and organs including roots, mature leaves, shoot apices and lateral buds. In contrast, the CsVMV promoter and its double enhancer derivative induced relatively weak expression in these tissues. Our results suggest that activity of the CsVMV promoter, in contrast to the CaMV 35S promoter, was under developmental regulation in transgenic grape plants as compared with the CaMV 35S promoter.
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Mid-Florida Research and Education Center, Institute of Food and Agricultural Sciences, University of Florida, 2725 Binion Road, 32703, Apopka, FL, USA
Jayasankar
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Gray
D J.
DJ