Abstract
The utility and commercial potential of genetically engineered (GE) plants would benefit from the use of site-specific recombination systems that enable efficient excision of the marker genes used to identify transformants. Although wheat is one of the most important food crops in the world, GE varieties have yet to be put into commercial production. To develop the Bxb1 recombination system (derived from the Mycobacterium smegmati bacteriophage Bxb1) for site-specific marker gene removal in transgenic wheat, we used biolistics to introduce into the wheat genome a codon optimized Bxb1 recombinase gene (BxbNom) under the control of the maize ubiquitin-1 promoter (Ubi1). Recombinase activity was monitored using a GUSPlus reporter gene activation assay. BxbNom recombinase-mediated excision of an att site-flanked stuffer DNA fragment activated β-glucuronidase reporter activity in callus, endosperm, and leaves in transient assays. The system also detected activity in leaves and endosperm of progeny of multiple independent transgenic wheat lines stably expressing BxbNom. Our results demonstrate that the Bxb1 recombinase is heritable in transgenic wheat plants and performs site-specific excision, providing a useful tool for generating marker-free GE plants. Establishment of wheat lines capable of efficiently excising unneeded marker genes removes one potential barrier to commercial deployment of GE wheat.
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Abbreviations
- GUSPlus :
-
Staphylococcus sp. β-glucuronidase gene
- eGFP:
-
Enhanced green fluorescent protein
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Acknowledgments
We thank Dr. V. Srivastava for seeds of the Cre-expressing transgenic wheat line ‘Cre37’ and Mara Guttman for excellent technical assistance. Mention of trade names or commercial products in this publication is solely to provide specific information and does not imply recommendation or endorsement by the U.S. Department of Agriculture. USDA is an equal opportunity employer.
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Blechl, A., Lin, J., Shao, M. et al. The Bxb1 Recombinase Mediates Site-Specific Deletion in Transgenic Wheat. Plant Mol Biol Rep 30, 1357–1366 (2012). https://doi.org/10.1007/s11105-012-0454-2
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DOI: https://doi.org/10.1007/s11105-012-0454-2