Genetic Manipulation - Environmental Biotechnology: Theory and Application - page 225

Resistance to herbicides
'Glyphosate', one of the most widely used herbicides, is an analogue of phos-
phoenol pyruvate and shows herbicidal activity because it inhibits the enzyme
5-enolpyruvylshikimate-3-phosphate synthase . The gene coding for this enzyme
has been identified, isolated and inserted into a number of plants including
petunias. In this case, the gene was expressed behind a CaMV promoter and intro-
duced using A. tumefaciens , leading to very high levels of enzyme expression.
As a consequence, the recombinant plants showed significant resistance to the
effects of glyphosate (Shah et al ., 1986). Developments in this strategy include
the formation of a chimaeric synthase enzyme, the analysis of which should lead
to improved herbicide resistance in transgenic crops using this strategy (He et al .,
2001).
An alternative approach but still using A. tumefaciens , has 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,
chlortoluron and chlorsulphuron (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 the -endotoxin of the bacterium, Bacillus thuringiensis , described

a little more fully in Chapter 10, have been transferred into plants. Examples
are of a synthetic B. thuringiensis

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- endotoxin gene transferred, in the first
case, by A. tumefaciens into Chinese cabbage (Cho et al ., 2001) and in the second,
by biolistic bombardment into maize (Koziel et 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 Magg et al . (2001). Insects are able to develop resistance
to Bt products which is a problem addressed by insertion of δ - endotoxin genes
into the chloroplast genome rather than into that of the plant's nucleus, with
promising results (Kota et al ., 1999; Daniell and Moar, 2007).
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 thuringiensis tends 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

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