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GENOMICS, MOLECULAR GENETICS & BIOTECHNOLOGY

Efficient Deletion of Transgenic DNA from Complex Integration Locus of Rice Mediated by Cre/lox Recombination System

^a Department of Crop, Soil & Environmental Sciences, University of Arkansas, Fayetteville, AR 72701
^b Department of Crop, Soil & Environmental Sciences, and Department of Horticulture, University of Arkansas, Fayetteville, AR 72701

^* Corresponding author (vibhas{at}uark.edu)

Transgenic plants may contain several DNA elements, most notably the selectable marker genes, that are not needed after the selection of the transgenic line. Using site-specific recombination system, such as Cre/lox, targeted deletion of these so-called unneeded DNA elements can be achieved. To study the efficiency of Cre/lox-mediated deletion of transgenic DNA from complex locus generated by particle bombardment, we generated two transgenic rice (Oryza sativa L.) lines that contain several copies of lox-flanked ß-glucuronidase (GUS) gene (gusA) and unmarked hygromycin resistance (hpt) gene and crossed them with a Cre-expressing line. Excision of both gusA and hpt genes occurred in F₁ hybrids. Molecular data demonstrated that Cre/lox recombination was initiated randomly in F₁ plants leading to complete excision of all transgene fragments in somatic tissue. F₂ analysis suggested that efficient excision also occurred in germline, preventing the transfer of lox-flanked genes into most of the F₂ plants. Further, the excised DNA did not apparently re-integrate into the genome. Thus, Cre/lox-mediated DNA recombination in rice is highly suitable for the removal of the unneeded transgenic DNA.

Abbreviations: bp, base pair • GUS, ß-glucuronidase • gusA, ß-glucuronidase gene • npt, neomycin phosphotransferase gene • hpt, hygromycin phosphotransferase gene • PCR, polymerase chain reaction

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