emerging technology and prospects of genetic engineering ... · emerging technology and prospects...

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Emerging technology and prospects of genetic

engineering to increase food production and quality

Dr. Joe KuhlDept. of Plant, Soil, &

Entomological Sciences

September 12, 2012

Crop Yield

• Thomas Malthus (1798) concluded that agricultural production increased arithmetically, yet population increased geometrically.

• Historically, the increase in area cultivated could keep up with demand, not so much anymore. Yield (=edible dry mass/unit area), therefore needs to increase!

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Facts: most of the dietary dry weight (and protein) for man comes from:

• five cereals

• three tuber crops

• several legumes

• two sugar crops

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GMO = Genetically Modified Organism

Also genetically modified (GM) or transgenic or genetically engineered (GE)

Gruskin 2012

Marshall 2012

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Marshall 2012

Marshall 2012

GM Plants

• Herbicide Tolerance– Transgene provides tolerance to broad-

spectrum herbicides (Roundup, Liberty, …)

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GM Plants

• Bt Insect-Resistant Crops– “Bt” short for Bacillus thuringiensis, a soil

bacterium whose spores contain a crystalline (Cry) protein

– Cry breaks down in insect gut to release a toxin (delta-endotoxin) – toxic to some insects

Marshall 2012

U.S. Regulatory systems

• Institutional Biosafety Committee (IBC)

• U.S. Department of Agriculture - Animal and Plant Health Inspection Service (APHIS)

• U.S. Environmental Protection Agency (EPA)

• Department of Health and Human Services -Food and Drug Administration (FDA)

• International agreements

http://www.colostate.edu/programs/lifesciences/TransgenicCrops/index.html

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What are GM plants?

Where do GM plants come from?

How are GM crops different?

What types of modifications are possible?

How could increased yield be achieved?

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Papaya Ring Spot Virus (PRSV)

• In 1992 PRSV was discovered in the Punadistrict of the Hawaiian Island (95% of the state’s papaya was grown)

PRSV – traditional control strategies

1) PRSV is efficiently spread plant to plant by ~60 species of aphids, non-persistent

2) No known natural resistance to the disease

- some tolerance, but this is rapidly lost

3) Use of netting is very expensive

4) Cross-protection is strain specific and not complete

5) Considerable variation exists in various strains

Genetically Engineered Resistance

• Genetically engineered plants contain the PRSV coat protein

• The coat protein is used by the plant to fight against the pathogen

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Questions to ask

• What plant species is modified?

• What gene or genes have been transferred?– Where does the gene(s) come from?

– How does the gene function?

• Where is the gene expressed, what tissue and when?

• What trait has been modified?

• How is management of the crop effected?

Modification Methods• Gene(s) Transfer

– Integration of “foreign” gene(s) into the plant genome

Trait

Organisms• Plants

• Animal

• Microbes

Genes

Traditional Breeding

Genetic Engineering

• Plants

• Animal

• Microbes

Organisms• Plants

• Animal

• Microbes

Genes• Plants

• Animal

• Microbes

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Genetic Engineering

• Transgenic CropsA transgenic crop plant contains a gene or genes

which have been “artificially” inserted instead of the plant acquiring them through pollination.

Method of introduction is key to definition, it does not matter what the resulting plant/product is.

Genetic Engineering

• TransgeneThe inserted gene sequence may come from

related or unrelated plant, or from a completely different species.

Example: transgenic papaya produces the PRSV coat protein

Example: transgenic Bt corn contains a gene from a bacterium

Genetics - DNA

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Modification Methods

• Gene(s) Transfer - Mechanisms– Physical

– Biological

– Alternative

• Gene modification

• Gene silencing


• Gene(s) Transfer– Physical

• Bombardment

• Microprojectile-mediated


• Gene(s) Transfer– Biological

• Agrobacterium tumefaciensmediated T-DNA transfer

Crown Gall

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• Gene(s) Transfer/Modification– Alternative methods:

• PEG-mediated protoplast transformation

• Electroporation mediated protoplast transformation

• Silicon carbide fiber

• Microinjections

• Desiccation (DNA uptake by dried embryos during rehydration)

• Targeted Gene Modification (TagMo)– Zinc-finger nucleases, Meganuclases, TALE nucleases,

DNA repair through homologous recombination

Kuzma & Kokotovich 2011

Gene Silencing

• Exploits plant regulatory mechanism– RNA Interference (RNAi)

• Targets specific plant gene(s)

• Decrease or eliminate expression

• May use siRNA or miRNA

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Silencing of Polyphenol Oxidase

Rommens et al. 2004



• Gene(s) Transfer – Variables:– What does the gene encode for?

– What organism is the gene from?

– What is the promoter?

– What is the desired phenotype?• Over-express, under-express

– Silencing: reduced or elimination of endogenous gene expression

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• Gene(s) Transfer – Variables

Variables (for each “event”)– Copy number

– Location in the plant cell

– Location in the plant genome

– Content of transferred genetic information

– Resulting phenotype

Organisms

Trait

Organisms

• Gene(s) Transfer – Plant Breeding and Testing

• Desired trait(s)– Activity of the introduced gene

– Stable inheritance of the gene

– Avoid unintended effects on plant growth, yield, and quality (off-types)

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Phenotypic Changes

• Off-types: genotype (cultivar) dependent– Extensive field testing

• Generate sufficient material and trial under field conditions– “Generally, majority of transgenic material

was phenotypically indistinguishable from control plants, and stable over several generations” (S. Millam)

GM Plants

Target Areas for Improvement:1. Biotic/abiotic stress resistance/tolerance and

yield

2. Biomass feedstock for biofuels

3. Value-added: nutrition, food functionality, quality traits

4. Plant production factories – therapeutics and industrial products

Potato - GM Traits

• Disease and pest resistance– Colorado potato beetle (cryIIIA)

– Potato tuber moth (cryV, cryI Ac9)

– Potato cyst nematodes (chicken egg white cystatin)

– Viruses, e.g. PLRV and PVY (sense and antisense)

– Bacteria and fungi• Erwinia and Phytophthora infestans

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Late Blight Resistance

Song et al. 2003

• Katahdin transformed with RB

Potato - GM Traits

• Tuber Quality– Anti-bruise (down-regulate PPO)

– Reducing sugars (over-express ADPglucosepyrophosphorylase; silence/reduce acid invertase)

Potato - GM Traits

• Gene silencing of vacuolar acid invertaseusing RNAi

Bhaskar et al. 2010

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Potato - GM Traits

• Nutritional value– Inulin (express artichoke genes)

– Carotenoids (down-regulate zeaxanthinepoxidase, express Erwinia phytoene synthase)

– Reduced glycoalkaloid content (down-regulate Stg1)

Potato - GM Traits

• Pharmaceutical– Vaccines, drug production

• Industrial– Starch

• High amylopectin and high amylose

Evaluating the use of GM plants

Consider GM plants on a case-by-case basis, taking into account:– Nature of the introduced gene

– Plant parts and growth stage in which the gene product is present

– Toxicity or allergenicity

– Presence of related wild species in the vicinity


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Evaluating the use of GM plants

Consider the environmental effects of alternatives to transgenic varieties:– Would more harmful chemicals be applied?

– Would conservation tillage change?

– What are the probability and effects of gene escape? Are wild relatives in the region?


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