Multigene Transformation Through Cre-lox Mediated Site-Specific Integration in Rice.

Plant transformation with multiple genes is a major challenge, rendering multi-trait engineering extremely difficult in crop plants. One of the hurdles in multigene transformation is the uncontrolled integration process that leads to low quality transgenic lines that are unsuitable for practical application. Recombinase-mediated site-specific integration has been tested and validated for developing high quality transgenic lines expressing one, two, or multiple genes. Of the numerous recombinase systems tested, Cre-lox and FLP-FRT show high efficiency in plants. Recently, Cre-lox system was successfully used to stack a set of 3 constitutive, 1 heat-induced, and 1 cold-induced gene. A number of transgenic lines were obtained through a relatively small effort, and the resulting transgenic lines all expressed the genes properly as determined by their promoter-specificity. Here, a method of Cre-lox mediated stacking of a multigene construct is described using rice as a model crop.