'GM-gene-deletor': fused loxP-FRT recognition sequences dramatically improve the efficiency of FLP or CRE recombinase on transgene excision from pollen and seed of tobacco plants

Abstract

Pollen- and seed-mediated transgene flow is a concern in plant biotechnology. We report here a highly efficient 'genetically modified (GM)-gene-deletor' system to remove all functional transgenes from pollen, seed or both. With the three pollen- and/or seed-specific gene promoters tested, the phage CRE/loxP or yeast FLP/FRT system alone was inefficient in excising transgenes from tobacco pollen and/or seed, with no transgenic event having 100% efficiency. When loxP-FRT fusion sequences were used as recognition sites, simultaneous expression of both FLP and CRE reduced the average excision efficiency, but the expression of FLP or CRE alone increased the average excision efficiency, with many transgenic events being 100% efficient based on more than 25,000 T(1) progeny examined per event. The 'GM-gene-deletor' reported here may be used to produce 'non-transgenic' pollen and/or seed from transgenic plants and to provide a bioconfinement tool for transgenic crops and perennials, with special applicability towards vegetatively propagated plants and trees.