Genetic Engineering of Canola

Introduction

Biotechnology is an all-encompassing term that describes the use of any living thing
or its parts (e.g. proteins, DNA, fungus, or bacteria) to make a product or to run a
process. Examples of traditional biotechnology include using yeast to make bread or
using bacteria to make yogurt. It also covers the bacterial and fungal inoculants that
farmers now use to provide nitrogen and phosphorus to crops.

The area of biotechnology that has been the focus of much attention and
controversy is genetic engineering. This technology involves taking DNA from one
organism and splicing it into the DNA of another organism. For herbicide resistance,
genetic material from common soil bacteria was introduced into canola. Varieties
using the Liberty Link and Roundup Ready systems were created this way. Another
system based on the herbicide bromoxynil is also in the works. Improvement on
“good fat” content was also taking place. This assignment will discuss on process,
problems, and progress involving genetically modified canola.

The Starting Point

Hundreds of years ago, Asians and Europeans used rapeseed oil in lamps. As time
progressed, people employed it as cooking oil and added it to foods. Its use was
limited until the development of steam power, when machinists found rapeseed oil
clung to water or steam-washed metal surfaces better than other lubricants. High
demand for rapeseed oil as a lubricant for the rapidly increasing number of steam
engines in naval and merchant ships happens during World War II. When the war
blocked European and Asian sources of rapeseed oil, a critical shortage developed
and Canada began to expand its limited rapeseed production.

After the war, demand declined sharply and farmers began to look for other uses for
the plant and its products. Edible rapeseed oil extracts were first put on the market
in 1956-1957, but these suffered from several unacceptable characteristics.
Rapeseed oil had a distinctive taste and a disagreeable greenish colour due to the
presence of chlorophyll. It also contained a high concentration of erucic acid.

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Experiments on animals have pointed to the possibility that erucic acid if consumed
in large quantities, may cause heart damage (Bellenand et al., 1980), although Indian
researchers Achaya K.T. (1987) have published findings that call into question these
conclusions and the implication that the consumption of mustard or rapeseed oil is
dangerous. Feed meal from the rapeseed plant was not particularly appealing to
livestock, due to high levels of sharp-tasting compounds called glucosinolates.

Plant breeders in Canada, where rapeseed had been grown (mainly in Saskatchewan)
since 1936, worked to improve the quality of the plant. In 1968, Dr Baldur Stefansson
of the University of Manitoba used selective breeding of two cultivars of rapeseed or
Brassica campestris (Brassica napus and B. campestris) (Brown et al., 1996) to
develop a variety of rapeseed low in erucic acid. In 1974, another variety was
produced low in both erucic acid and glucosinolates; it was named canola, from
Canadian oil, low acid (Stoorgard, 2008). Their seeds are used to produce edible oil
that is fit for human consumption because it has lower levels of erucic acid than
traditional rapeseed oils and to produce livestock feed because it has reduced levels
of the toxin glucosinolates.

The Process

Each canola plant produces yellow flowers that, in turn, produce pods; similar in
shape to pea pods about one-fifth the size. Within the pods are tiny round seeds that
are crushed to obtain canola oil. Each seed contains approximately 40 per cent oil.

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Genetically-modified canola
Genetically modified (GM) or transgenic canola varieties made by utilizing modern
plant breeding or genetic engineering have been developed to be resistant to
specific herbicides. They are called herbicide-resistant varieties. The plants are
modified, but the oil is not modified.

Agriculture giant, Monsanto introduced GM canola variety GT73 that are herbicide-
resistant in 1997. It was produced by inserting the EPSPS gene encoding the enzyme
5-enolypyruvylshikimate-3-phosphate synthase (EPSPS) from the CP4 strain of
Agrobacterium tumefaciens and glyphosate oxidase (gox) gene isolated from strain
LBAA Ochrobactrum anthropi. Agrobacterium-mediated transformation method was
used, utilizing single insertion event containing one copy of the T-DNA from plasmid
PV-BNGT04 (Batista and Oliviera, 2009). The T-DNA consists of CmoVb promoter
(modified figwort mosaic virus), the CP4 EPSPS gene, the gene coding GOX v247 and
the E9 3' polyadenylation signals. Only the DNA required to confer the glyphosate-
tolerance phenotype was transferred and inserted at a single locus in the canola
genome. No genetic elements from outside of the right and left borders of the
plasmid were transferred into or are present in the genomic DNA of Roundup Ready
canola.

The EPSPS gene codes for the enzyme 5-enolpyruvylshikimate-3-phosphate synthase

that is present in all plants, bacteria and fungi. The EPSPS enzyme is part of an

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important biochemical pathway in plants called the shikimate pathway, which is

involved in the production of aromatic amino acids and other aromatic compounds.
When conventional canola plants are treated with glyphosate, the plants cannot
produce the aromatic amino acids needed to grow and survive. EPSPS is not present
in mammals, birds or aquatic life forms, which do not synthesize their own aromatic
amino acids. For this reason, glyphosate has little toxicity to these organisms. The
EPSPS enzyme is naturally present in foods derived from plant and microbial sources.
Glyphosate oxidase (gox) enzyme accelerates the normal breakdown of the herbicide
glyphosate into two non-toxic compounds, aminomethylphosphonic acid (AMPA)
and glyoxylate. AMPA is the principal breakdown product of glyphosate and is
degraded by several microorganisms, while glyoxylate is commonly found in plant
cells and is broken down by the glyoxylic pathway for lipid metabolism.

Plasmid map for PV-BNGT04

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Main stages required for production of GM canola plants: EPSPS gene and gox gene
isolated from donor organism (a), cloned in vector plasmid PV-BNGT04 with CmoVb
as promoter and E9 3' polyadenylation signals (b) and transferred into the receptor
organism (c). The receptor plant cells are then selected and regenerated to obtain
complete GM plants (d). Finally, the obtained GE plants are crossed with other,
already improved plants (e). The regenerated plants and their progenies are
analysed at various levels throughout this process.

Australian scientist J.M. Manners and K. Kazan are undertaking studies aimed at the
development of transgenic canola plants expressing novel genes for resistance to
fungal pathogens such as Leptosphaeria maculans, Sclerotinia sclerotiorum,
Rhizoctonia solani and Alternaria brassicicola. The genes being tested include those
encoding novel antimicrobial peptides derived from Australian native plants and
from macadamia in particular. So far, transgenic plants expressing three peptides
have been developed and advanced lines expressing one peptide at high levels are
under evaluation for improved disease resistance. Other related research aimed at
the development of transgenic canola plants, expressing genes encoding antifungal
proteins from Brassicaceae seeds, for resistance to L. maculans and S. sclerotiorum.
Transgenic approaches employed to enhance yield and to develop novel inducible
male sterility systems for hybrid seed production.

Varieties with very high oleic acid types was also developed based on elite Australian
B. napus breeding lines and advanced double-low B. juncea germplasm.

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Modifications to existing Brassica transformation protocols and the use of an intron-

interrupted hygromycin-resistance gene as the selectable marker have resulted in
improved transformation efficiencies. Silencing of the endogenous oleate desaturase
genes have resulted in substantial increases in oleic acid levels, up to 89% in B. napus
and 73% in B. juncea (Graef et al., 2007).

Current research also involves development of canola with new trait such as
photoperiod insensitivity, modified plant architecture, and with reduced yield loss.
Various modern genetic engineering methods have been employed such as
protoplast fusion and microspore technologies. Embryo rescue method used to
widen genetic base of rapeseed by hybridization of B. oleracea and B. campestris to
resynthesize B. napus (Inomata, 1978).

Benefits

Compared to conventional selection and breeding of living organisms, the

combination of genetics, molecular biology, and cell biology that underlies genetic
engineering of plants allows for: reduced time to achieve genetic improvement;
more precise genetic manipulation (a single gene or set of genes of known function
can be transferred from one organism to another); and a wider range of genetic
changes, as the genetic transfers facilitated via genetic engineering are often ones
that would not normally occur in nature.

In conventional canola, there are some weeds that are difficult to control such as
wild mustard, stinkweed, cleavers and shepherd's purse. Growers choose herbicide-
resistant canola varieties primarily because weed control is easier, cost effective (up
to 30% saving on pesticides), and better (Brown et al., 1996). They allow farmers to
produce canola on land where it was previously hard to get good yields because of
high broadleaf weed populations. These varieties have also been effective at
reducing dockage in canola, and at reducing future weed problems and pesticide use.
Crop establishment and low plant populations are often problems with canola
because of soil crusting and poor emergence. Herbicide-tolerant varieties provide

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good weed control and allow producers to leave thinner crop stands to still obtain
acceptable yields. Reseeding can be avoided and the farmer’s bottom line is
improved. Herbicide resistant canola can be used in direct seeding systems to reduce
erosion and lower the carbon content in the air. Nearly 90% of canola cultivation at
Canada is herbicide-tolerant (Brown et al., 1996).

Genetic engineering is also being directed toward products that benefit consumers:
more nutritious foods, foods with medicinal value, and foods with improved cooking
characteristics. Canola oil has been claimed to promote good health due to its very
low saturated fat and high monounsaturated fat content, and beneficial omega-3
fatty acid profile. The Canola Council of Canada states that it is completely safe and
is the "healthiest" of all commonly used cooking oils. It has well established heart
health benefits and is recognized by many health professional organizations
including the American Dietetic Association and American Heart Association (Lorgeril
and Salen, 2006). Canola oil has been authorized a qualified health claim from the US
Food and Drug Administration based on its ability to reduce the risk of coronary
heart disease due to its unsaturated fat content.

Concerns and Challenges

Various studies confirm that outcrossing, or gene transfer, does happen, albeit at a
rate of less than 0.25% in a typical 160 acre field. Research has shown that most of
the outcrossing happens within 100 metres of the field edge (Kott et al., 1990; Linder,
1994). Crawley et al. (1993), who staged one of the largest-scale plant population
studies of all time, to evaluate the invasiveness of conventional canola and canola
genetically engineered to express resistance to the antibiotic kanamycin or the
herbicide glufosinate. Crawley et al. measured the finite rate of increase of these
lines of canola in four habitat types in each of three sites, with a barrage of
treatments that included cultivation/no cultivation, fencing, molluscicide, insecticide,
and fungicide. They found that only in cultivated plots can canola replace itself, and
in no case did the transgenic plants exhibit a higher finite rate of increase than the
conventional plants.

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Canola Council of Canada refers this concern as isolated case and urging the farmers
to develop herbicide use pattern to manage the problem of outcrossing when
different types of canola, such as Roundup Ready, Liberty Link, Smart or
conventional varieties are grown side by side. There are many broadleaf herbicides
on the market that will control this problem. The familiar 2,4-D or MCPA, used in
rotational crops such as cereals, are effective.

GM canola facing similar challenges like other GM crops such as corn, cotton, and
soybean due to strong objections about some aspects of genetic engineering.
Opponents range from those who believe the techniques should not be used at all to
those who believe products based on the techniques should be labelled as such. The
fierce debate continues even though supporters of genetic engineering have
tempered, and in some cases refuted, the arguments advanced by the opposition.

Those who oppose genetic engineering argue that these techniques are largely
untested. They assert that genetically engineered crops and foods are not
“substantially equivalent” to those produced using traditional means and should not
be expected to have similar health and environmental impacts. Among the major
health concerns is the potential for creating allergenic properties in foods. One
widely reported case involved the development of a genetically transformed
soybean variety that, in the course of field trials, was found to contribute to allergies
(Bailey, 2000). The long-term environmental impacts of growing genetically
engineered canola are also being questioned (Weick and Walchli, 2002)

Genetic engineering has also been opposed on ethical and religious grounds because
the techniques allow for cross-fertilizing that would not occur in nature (Moseley,
1999). The role of business - particularly large multinationals - in the research,
development, and marketing of genetically engineered crops and foods also has
caused some scepticism. Critics argue that agricultural biotechnology is dominated
by large companies that may allow profits to blind them to potential risks. Some are
concerned that these companies will tend to favour applications with large markets

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where the products can be afforded, i.e. mostly in industrialized countries - rather
than farmers with smaller holdings in developing countries. Others fear that
developing countries will be used as a “dumping ground” for untested technology. In
fact, the very need for greatly increased amounts of food provided by genetic
engineering has been questioned. The world food shortage, opponents argue, is not
a matter of quantity but of distribution and affordability, and they argue that these
problems will not be solved via genetic engineering

The Future

Canola is the workhorse of the agricultural biotechnology industry. Compared to

other crops, its DNA is relatively easy to modify. Canola can be engineered to
produce medicines, or to produce a specific fatty acid for speciality markets or
industrial uses. The quality of the oil and protein can be improved, to produce better
products for frying and salad oil, or high quality livestock feed. Plant biotechnology is
adding value to canola. In this first generation of genetically modified crops, the
benefits have largely gone to the seed companies and producers. In the future,
consumers will see direct benefits as well.

References
Achaya, K.T. 1987. Fat Status Among Indians - A Review. Journal of Scientific and
Industrial Research. 46: 112-126.

Bailey P. 2000. Promise or Peril? Are Genetically Engineered Foods Cause for
Concern? UC Davis Magazine. 23(5): 23-26.

Batista, R. and Oliviera, M.M. 2009. Fact or Fiction of Genetically Modified Food.
Trends in Biotechnology. 27(5): 277-286.

Bellenand, J.F., Baloutch, G., Ong, N., and Lecerf, J. 1980. Effects of Coconut Oil on
Heart Lipids and on Fatty Acid Utilization in Rapeseed Oil. Lipids. 15(11): 938-
943.

Brown, J, Thill, D.C., Brown, A.P., Mallory-Smith, C., Brammer, T.A., and Nair, H.S.
1996. Gene Transfer Between Canola (Brassica napus) and Related Weed
Species. Annals of Applied Biology. 129(3): 513-522.

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Crawley, M.J., Hails, R.S., Rees, M., Kohn, D., and Buxton, J. 1993. Ecology of
Transgenic Oilseed Rape in Natural Habitats. Nature. 363: 620-623.

Graef, F., Stachow, U., Werner, A., and Schutte, G. 2007. Agricultural Practice
Changes with Cultivating Genetically Modified Herbicide-tolerant Oilseed
Rape. Agricultural Systems. 94(2): 111-118

Inomata, N. 1978. Production of Interspecific Hybrids of B. campestris x B. oleracea

by Culture In Vitro of Ovaries. Japan Journal of Genetics. 53: 161-173.

Kott, L.S., Erickson, L.R., Beversdorf, W.D. 1990. The Role of Biotechnology in
Canola/Rapeseed Research. In Shahidi, F. (ed), Canola and Rapeseed:
Production, Chemistry, Nutrition, and Processing Technology. pp 47-80. Van
Nostrand Reinhold, New York, USA.

Linder, C. R. 1994. The Ecology of Population Persistence for Wild, Crop, and Crop-
wild Hybrid Brassica and its Implications for Transgenes Escaped from Canola.
PhD Thesis. Brown University, Providence, USA.

Lorgeril, M, and Salen, P. 2006. The Mediterranean-style Diet for the Prevention of
Cardiovascular Diseases. Public Health Nutrition. 19(5): 118-123.

Moseley, B.E.B. 1999. The Safety and Social Acceptance of Novel Foods. International
Journal of Food Microbiology. 50(6): 25-31.

Salisbury, P. 1998. Genetically Modified Oilseeds – The Impact of Gene Technology on

the Australian Oilseeds Industry. Australian Oilseeds Federation Innovations
and Technology Committee, Sydney, Australia.

Storgaard, A.R. 2008. Stefansson, Baldur Rosmund. In The Editors, The Canadian
Encyclopedia, pp 600-602. Hurtig Publisher, Edmonton, Canada.

Weick, C.W. and Walchli, S.B. 2002. Genetically Engineered Crops and Foods: Back to
the Basics of Technology Diffusion. Technology in Society. 24(3): 265-283

http://dictionary.infoplease.com/canola (131009)

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