Asexual production of selectable marker-free transgenic woody plants, vegetatively propagated species |
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Authors: | Etsuko Matsunaga Koichi Sugita Hiroyasu Ebinuma |
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Affiliation: | (1) Wood-Bioengineering, Pulp and Paper Research Laboratory, Nippon Paper Industries Co., Ltd., 5-21-1 Oji, Kita-ku, Tokyo, 114-0002, Japan |
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Abstract: | We describe here a practical system for generating selectable marker-free transgenic woody plants independent of sexual crossing. We previously reported that the GST-MAT vector system could produce marker-free transgenic tobacco plants containing a single-copy transgene at high frequency. The GST-MAT vector system consists of a DNA excision cassette of the R/RS site-specific recombination system from Zygosaccharomyces rouxii into which the isopentenyltransferase gene from Agrobacterium tumefaciens has been inserted. In this study, we applied this new GST-MAT vector to hybrid aspen (Populus Sieboldii X Populus grandidentata), a model of vegetatively propagated plant species, to produce selectable marker-free transgenic woody plants. In the new GST-MAT vector, the chimeric ipt gene fused with a light-inducible rbcS promoter efficiently produced transgenic ipt-shooty with GUS activity from 38.0% of infected stems. Upon excision of the R and ipt genes between RS sites, regulated by the inducible promoter of the maize glutathione-S-transferase (GST-II-27) gene, 3 (21.4%) transgenic hybrid aspen plants with marker-free and normal phenotype were generated from 14 ipt-shooty lines within 2 months after cutting induction. These results suggest that the MAT-vector system might be useful for removing a selectable marker gene independent of sexual crossing in vegetatively propagated species. |
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Keywords: | Aspen Glutathione-S-transferase Isopentenyltransferase Marker-free transgenic Site-specific recombination Transformation |
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