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Effective selection system for generating marker-free transgenic plants independent of sexual crossing. Plant Cell Reports 18, 941-947.
| Sugita K., Matsunaga, E., Ebinuma, H. (1999).
| In a previous report, a novel selection protocol termed "the MAT-vector system" for generating marker-free transgenic plants (MFTPs) was presented. The first stage of the system is visual selection of morphologically abnormal transgenic shoots, ipt-shooty, that have lost apical dominance and rooting ability. The second stage involves elimination of the ipt gene and the appearance of MFTPs free of ipt gene influence. The present report describes a practical MAT-vector in which removal of the ipt gene is efficiently mediated by the site-specific recombination system R/RS from Zygosaccharomyces rouxii, in place of the maize transposable element Ac, used previously. This improved MAT-vector produced MFTPs from 39% of moderate ipt-shooty and 70% of extreme ipt-shooty lines. These results are superior to the previous MAT-vector which produced MFTPs from only 5% of ipt-shooty lines. The present novel system also induced direct development of MFTPs from adventitious buds without production of ipt-shooty intermediates. The presence of beta-glucuronidase (GUS) and neomycin phosphotransferase (NPTII) genes of interest, and the absence of the ipt gene were verified by a GUS histochemical assay, NPTII assay, and molecular analysis.
A novel selection protocol termed " the MAT-vector system" for producing marker -free transgenic plants, which involves visual selection of morphologically abnormal transgenic shoots and the following elimination of the ipt gene, was presented previously. Here the removal of the ipt gene is efficiantly mediated by the site-specific recombination system R/RS from Zygosaccharomyces rouxii, instead of the maize Ac system used before. The improved systems produced marker free transgenic plants from 39% of moderate ipt-shooty and 70% of extreme ipt-shooty lines, as compared to the 5% reported with the previous system.
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