• Open Access

Site-specific gene integration in rice genome mediated by the FLP–FRT recombination system

Authors

  • Soumen Nandy,

    1. Department of Crop, Soil & Environmental Sciences, University of Arkansas, Fayetteville, AR, USA
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  • Vibha Srivastava

    Corresponding author
    1. Department of Crop, Soil & Environmental Sciences, University of Arkansas, Fayetteville, AR, USA
    2. Department of Horticulture, University of Arkansas, Fayetteville, AR, USA
      Correspondence (Tel: 479 575 4872; fax: 479 575 7465; email vibhas@uark.edu)
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Correspondence (Tel: 479 575 4872; fax: 479 575 7465; email vibhas@uark.edu)

Summary

Plant transformation based on random integration of foreign DNA often generates complex integration structures. Precision in the integration process is necessary to ensure the formation of full-length, single-copy integration. Site-specific recombination systems are versatile tools for precise genomic manipulations such as DNA excision, inversion or integration. The yeast FLP–FRT recombination system has been widely used for DNA excision in higher plants. Here, we report the use of FLP–FRT system for efficient targeting of foreign gene into the engineered genomic site in rice. The transgene vector containing a pair of directly oriented FRT sites was introduced by particle bombardment into the cells containing the target locus. FLP activity generated by the co-bombarded FLP gene efficiently separated the transgene construct from the vector-backbone and integrated the backbone-free construct into the target site. Strong FLP activity, derived from the enhanced FLP protein, FLPe, was important for the successful site-specific integration (SSI). The majority of the transgenic events contained a precise integration and expressed the transgene. Interestingly, each transgenic event lacked the co-bombarded FLPe gene, suggesting reversion of the integration structure in the presence of the constitutive FLPe expression. Progeny of the precise transgenic lines inherited the stable SSI locus and expressed the transgene. This work demonstrates the application of FLP–FRT system for site-specific gene integration in plants using rice as a model.

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