Genetic Manipulation 227petunias. In this case, the gene was expressed behind a CaMV promoter and
introduced usingA. tumefaciens, leading to very high levels of enzyme expres-
sion. As a consequence, the recombinant plants showed significant resistance to
the effects of glyphosate (Shahet al. 1986). Developments in this strategy include
the formation of a chimaericsynthaseenzyme, the analysis of which should lead
to improved herbicide resistance in transgenic crops using this strategy (He 2001).
An alternative approach but still usingA. tumefacienshas been to transfer the
genes for mammalian cytochrome P450 monooxygenases, known to be involved
in the detoxification (and activation) of many xenobiotics including pesticides,
into tobacco plants. These transgenics displayed resistance to two herbicides,
chlortoluronandchlorsulphuron(Yordanova, Gorinova and Atanassov 2001).
Improved resistance to pests
Plants have an inbuilt defence mechanism protecting them from attack by insects
but the damage caused by the pests may still be sufficient to reduce the com-
mercial potential of the crop. The usual procedure is to spray the crop with
insecticides but in an effort to reduce the amount of chemical insecticides being
used, plants are being engineered to have an increased self-defence against pests.
Attack by insects not only causes damage to the plant but also provides a route
for bacterial or fungal infection in addition to the role played in the spread of
plant viruses. With a view to increasing resistance to sustained attack, the genes
coding for theδ-endotoxinof the bacterium,Bacillus thuringiensis(Bt), described
a little more fully in Chapter 10, have been transferred into plants. Examples are
of syntheticB. thuringiensisδ-endotoxingenes transferred, in the first case, by
A. tumefaciensinto Chinese cabbage (Choet al. 2001) and in the second, by
biolistic bombardment into maize (Kozielet al. 1993). In both cases, the trans-
genic plants showed greatly improved resistance to pest infestation. There are,
however, some problems with crop performance of some genetically engineered
plants highlighted in Magget al. (2001). Insects are able to develop resistance
to Bt products which is a problem addressed by insertion ofδ-endotoxingenes
into the chloroplast genome rather than into that of the plant’s nucleus, with
promising early results (Kotaet al. 1999).
It may be recalled, that for each amino acid incorporated into a protein there
is usually a choice of three or four codons all of which code for that same amino
acid. Different organisms have distinct preferences for a particular codon, thus
Bacillus thuringiensistends to use codons richer in thymidine and adenine than
the plant cells into which the gene is placed. There are also signals controlling the
expression of these genes relevant to bacteria, rather than eukaryotes, which will
not function very well, if at all, in the plant cell. For these reasons, expression
may benefit from modification of the DNA sequence to compensate for these
differences while maintaining the information and instructions. This may account
in part for the very high levels of expression and stability of the Bt proteins whose
genes have been introduced, by (biolistic) microbombardment, into chloroplasts