of sites within a genome could potentially lead to the large-scale restructuring of
chromosomes. Of these potential applications, the removal of DNA by site-specific recom-
bination has already been adopted for commercial product development. Renessen, a joint
venture between Cargill and Monsanto, introduced a maize line, LY038, for use in animal
feed, marketed under the trade name MaveraTM. High-value corn with lysine, LY038, is
derived from biolistic transformation of maize to incorporate acordapAgene that directs
seed-specific production of a lysine-insensitive dihydrodipicolinate synthase enzyme.
The kanamycin resistance marker used for transformation, nptII, was subsequently
removed by site-specific recombination, a process first described in model plants some
15 years earlier (Dale and Ow 1991; Russell et al. 1992). In the transformation vector,
nptIIwas flanked by directly orientedloxsites from the Cre-loxrecombination system
(Fig. 16.2). On recovery of the desired transformant, a cre gene was introduced
into the genome from a genetic cross. Cre recombinase excised away thelox-flanked
DNA, and thecregene was then removed by genetic segregation. With a decade-long
development timeline for genetically modified (GM) maize, the decision to incorporate a
recombinase-mediated marker removal step must have been made in the mid-1990s.
Another example is the ability to remove transgenes from pollen to delimit unwanted
gene flow to wild relatives and nontransgenic plants that are sexually compatible. Not
only are crop-wild transgenic hybrids undesirable from the perspective of transgene
escape; repeated backcrossing of transgenes into wild relatives is of regulatory and consu-
mer concern as well. This introgression of transgenes is already likely very rare (Stewart
et al. 2003), but could probably be eliminated entirely by a “GM-gene-deletor system”
(Luo et al. 2007). In such a system, transgenes of interest are located within dual fused
recombination recognition sites with the recombinase under the control of a pollen-specific
promoter. Only in pollen grains would the transgenes be deleted, and thus risks of hybrid-
ization and introgression would be greatly decreased (Fig. 16.3).
16.2.2 More Precise Integration of DNA
As commercial products improve over time through new innovations, it is likely that
additional features will be incorporated into future generations of transgenic plants and
Figure 16.2.Renessen’s high-lysine corn line LY038 used site-specific recombination to remove the
transformation selectable marker, the kanamycin resistance genenptII, after stable incorporation of
cordapAthat directs high-lysine production in seed. Cre recombinase, introduced from hybridization
with acretransgenic plant, excised thenptIImarker flanked by directly orientedloxrecombination
sites. Thecregene was subsequently segregated away in the following generation.
362 THE FUTURE OF PLANT BIOTECHNOLOGY