Principles of Plant Genetics and Breeding

Figure 3 Field trials demonstrating gene efficacy.
Figure 4 Selecting the commercial event.
BREEDING FOR PHYSIOLOGICAL AND MORPHOLOGICAL TRAITS 359
promoter (e35S), are expressed strongly in
wheat leaves, but at lower levels in specific
tissues critical for reproduction. Other promoters,
like rice actin, are expressed at
higher levels in these critical reproductive
tissues but at lower levels during plant
regeneration and in wheat leaves. In
wheat, we used both the e35S and rice
actin promoters to achieve plant selection
and a commercial level of vegetative and
reproductive tolerance to glyphosate.
Usually, many different constructs with
different combinations of promoters, genes,
and stop regions are tested to find the
combination that provides the desired
phenotypic result.
First, tests for the function of the introduced
gene – for example, in the case of
Roundup Ready® wheat, glyphosate spray
tests – are conducted to select transgenic
plants that express the trait. Seed of each
event must be increased in greenhouses
or growth chambers to provide seed for
field testing. Any “environmental release”
(field trial) is conducted under the rules
and regulations of the US Department
of Agriculture–Animal and Plant Health
Inspection Service (USDA-APHIS) as well
as individual state departments of agriculture.
Similarly, the Canadian Food Inspection
Agency (CFIA) controls field testing
in Canada. For wheat, these regulations
include specifications regarding minimum
isolation distances, volunteer monitoring
requirements, and crop rotation restrictions.
Field trials are conducted over several
locations and years to obtain enough performance
data to select the commercial
transgenic event (Figure 3) (Zhou et al. 2003).
Selecting the commercial transgenic
event
In order to facilitate the selection of the
eventual commercial event, it is most useful
for the team to set selection criteria before the data is analyzed. Selection criteria involves four areas: (i) molecular biology (single
insertion, no vector backbone, intact insertion); (ii) genetics (trait inherited as a single dominant gene, expression maintained
over generations); (iii) agronomics (yield, maturity, disease reactions – i.e., selection of plants that are not affected by the transformation/regeneration
process); and (iv) trait performance (for herbicide tolerance, at least a 2× safety margin to the commercial
rate and timing of application) (Figure 4). In the case of Roundup Ready® wheat, the development team invoked an additional
selection criterion to select a transgenic event that was present on the A or B genome. Molecular breeding techniques were used
to map the insertion of every potential commercial event. By selecting away from the D genome, research demonstrates that there
is a high likelihood that crossing between wheat and its only North American wild relative, jointed goatgrass (Aegilops cylindrica),
would not result in introgression of the trait into the wild jointed goatgrass population.
Developing the trait
In parallel, the push is often underway in four primary areas: (i) developing regulatory packages for the Environmental Protection