Efficient deletion of the selectable marker gene from transgenic citrus via the Cre/loxp system

Selectable marker genes conferring antibiotic- or herbicide- resistance are in general required to efficiently recover transgenic plants from transformed cells during plant transformation. However, the presence of marker genes raises public concerns regarding the field release of transgenic plants. Removal of marker genes prevents the risk of its release into the environment and hastens the public acceptance of transgenic products. In this study, an efficiency system for deletion of a selectable marker gene from transgenic citrus by Cre/loxp siterecombination has been constructed via conventional Agrobacterium-mediated transformation. Our results showed that about 20% of transformation efficiencies could be obtained using ipt (isopentenyltransferase) gene from A. tumefaciens as a selectable marker gene. Molecular analysis demonstrated that 20-100% complete deletion occurred in transgenic plants and that Cre/loxp-mediated excision was always precise in citrus. This approach provides a reliable strategy for deleting selectable marker genes from transgenic citrus after transformation and thus producing marker-free transgenic plants.