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The Science of Transgenics

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  1. The Science of Transgenics The Sociological, Economic, and Ethical Impact of Transgenic Organisms Workshop Fargo, ND February 21, 2003 Phil McClean Department of Plant Science North Dakota State University

  2. Transgenics are a Biotechnology Product How about some definitions Biotechnology - General Definition The application of technology to improve a biological organism Biotechnology - Detailed Definition The application of the technology to improve the biological function of an organism by adding genes from another organisms

  3. What About the Term Genetic Engineering? Genetic engineering is the basic tool set of biotechnology Genetic engineering involves: • Isolating genes • Modifying genes so they function better • Preparing genes to be inserted into a new species • Developing transgenes

  4. What is a transgenic? Concept Based on the Term Transgene Transgene – the genetically engineered gene added to a species Ex. – modified EPSP synthase gene (encodes a protein that functions even when plant is treated with Roundup) Transgenic – an organism containing a transgene introduced by technological (not breeding) methods Ex. – Roundup Ready Crops

  5. Why are transgenics important? We can develop organisms that express a “novel” trait not normally found in the species Extended shelf-life tomato (Flavr-Savr) Herbicide resistant soybean (Roundup Ready)

  6. Agriculture Transgenics On the Market • Insect resistant cotton – Bt toxin kills the • cotton boll worm • transgene = Bt protein Source: USDA • Insect resistant corn – Bt toxin kills the • European corn borer • transgene = Bt protein Normal Transgenic

  7. Herbicide resistant crops • Now: soybean, corn, canola • Coming: sugarbeet, lettuce, strawberry • alfalfa, potato, wheat (2005?) • transgene = modified EPSP synthase or • phosphinothricin-N-acetyltransferase Source: Monsanto • Virus resistance - papya resistant to • papaya ringspot virus • transgene = virus coat protein

  8. Biotech chymosin; the enzyme used • to curdle milk products • transgene = genetically engineered enzyme Source: Chr. Hansen • bST; bovin somatotropin; used to increase • milk production • transgene = genetically engineered enzyme Source: Rent Mother Nature

  9. Some Ag Biotech Products Are Discontinued Why??? • Poor Quality • FlavrSavr tomatoes (Calgene) • Negative Consumer Response • Tomato paste (Zeneca) • Negative Corporate Response • NewLeaf (Monsanto) • Universal Negative Publicity • StarLink corn (Aventis)

  10. Next Generation of Ag Biotech Products Golden Rice – increased Vitamin A content (but not without controversy) transgene = three pathway enzymes Sunflower – white mold resistance transgene = oxalate oxidase from wheat Source: Minnesota Microscopy Society

  11. Turfgrass – herbicide resistance; slower growing (= reduced mowing) Bio Steel – spider silk expressed in goats; used to make soft-body bullet proof vests (Nexia)

  12. Biotechnology is Not Just on the Farm Disease Treatment Diagnostics Environmental Cleanup Human Applications

  13. Human Applications • Pharmaceutical products • New solutions to old problems • Disease diagnosis • Determine what disease you have or may get  • Gene therapy • Correcting disease by introducing a corrective gene

  14. Biotechnology and Health The genes for these proteins are: • Cloned • Inserted into bacteria • Product isolated using biofermentation

  15. Environmental Applications Bioremediation - cleanup contaminated sites; uses microbes designed to degrade the pollution Indicator bacteria – contamination can be detected in the environment

  16. Future Health-related Biotech Products Vaccines – herpes, hepatitis C, AIDS, malaria Tooth decay – engineered Streptococcus mutans, the bacteria that destroys enamel

  17. Edible Vaccines Transgenic Plants Serving Human Health Needs • Works like any vaccine • A transgenic plant with a pathogen protein gene is developed • Potato, banana, and tomato are targets • Humans eat the plant • The body produces antibodies against pathogen protein • Humans are “immunized” against the pathogen • Examples: • Diarrhea • Hepatitis B • Measles

  18. A Popular Term We Need To Know GMOs - Genetically modified organisms • GMO - an organism that expresses traits that result • from the introduction of foreign DNA • Originally a term equivalent to transgenic organism

  19. The GMO Ruse • Some claim any improved biological product is a GMO • They feel this will • ease the publics fear • pave the way for product acceptance • For example, some call plant varieties biotechnology products • This is a false claim

  20. Let’s Be Up Front • Biotechnology adds traits not available in the species •  Soybean does not have a gene to breakdown Roundup • The gene comes from bacteria • Breeding  Biotechnology • Breeding only exchanges genes found in the species • Breeding can transfer the transgene to other breeding materials • BUT this does not make it a biotechnology procedure

  21. Important Plant Improvement Methods • Breeding • Crossing two individuals from the same species; • produces a new, improved variety; • not a biotechnology procedure Source: USDA • Transformation • Adding a gene from another species; the • essential biotechnology procedure to produce • transgenics Source: USDA

  22. Interspecific Cross Wheat Rye X Triticale New species, but NOT biotechnology products

  23. Mutagenesis • A useful procedure to produce a new trait • But the normal gene is modified • A transgene is not involved • The product of mutagenesis is not a GMO

  24. Mutagenesis Changes the DNA Sequence Mutagenesis Treatment Susceptible Normal Gene ATTCGA Resistant Mutant Gene ATTGGA

  25. BASF Clearfield Products • Herbicide resistance • imidazolinones • Mutant AHAS enzyme • developed by mutagenesis • Crops • Canola • Corn • Rice • Sunflower • Wheat • A Major Marketing Advantage • but lost when stacked with a transgene

  26. The Roundup Ready Story • Glyphosate is a broad-spectrum herbicide • Active ingredient in Roundup herbicide • Kills all plants it come in contact with • Inhibits a key enzyme (EPSP synthase) in an amino acid pathway • Plants die because they lack the key amino acids • A resistant EPSP synthase gene allows crops • to survive spraying

  27. Shikimic acid + Phosphoenol pyruvate Plant EPSP synthase 3-Enolpyruvyl shikimic acid-5-phosphate (EPSP) Aromatic amino acids Roundup Sensitive Plants + Glyphosate X X Without amino acids, plant dies X X

  28. Roundup Resistant Plants Shikimic acid + Phosphoenol pyruvate + Glyphosate RoundUp has no effect; enzyme is resistant to herbicide Bacterial EPSP synthase 3-enolpyruvyl shikimic acid-5-phosphate (EPSP) With amino acids, plant lives Aromatic amino acids

  29. The Golden Rice Story • Vitamin A deficiency is a major health problem • Causes blindness • Influences severity of diarrhea, measles • >100 million children suffer from the problem • For many countries, the infrastructure doesn’t exist • to deliver vitamin pills • Improved vitamin A content in widely consumed crops • an attractive alternative

  30. IPP Geranylgeranyl diphosphate Phytoene synthase Phytoene Problem: Rice lacks these enzymes Phytoene desaturase ξ-carotene desaturase Lycopene Lycopene-beta-cyclase Normal Vitamin A “Deficient” Rice  -carotene (vitamin A precursor) -Carotene Pathway Problem in Plants

  31. IPP Geranylgeranyl diphosphate Phytoene synthase Phytoene Vitamin A Pathway is complete and functional Phytoene desaturase ξ-carotene desaturase Lycopene Lycopene-beta-cyclase  -carotene (vitamin A precursor) Golden Rice The Golden Rice Solution -Carotene Pathway Genes Added Daffodil gene Single bacterial gene; performs both functions Daffodil gene

  32. Introducing the Gene or Developing Transgenics Steps 1. Create transformation cassette 2. Introduce and select for transformants

  33. 1. Gene of interest • The coding region and its controlling elements 2. Selectable marker • Distinguishes transformed/untransformed plants 3. Insertion sequences • Aids Agrobacterium insertion Transformation Cassettes Contains

  34. Tissue must be capable of developing into normal plants • Leaf, germinating seed, immature embryos • Develop shoots • Root the shoots Transformation Steps Prepare tissue for transformation Introduce DNA • Agrobacterium or gene gun Culture plant tissue Field test the plants • Multiple sites, multiple years

  35. Agrobacterium Tissue culture required to generate transgenic plants • Gene Gun Delivering the Gene to the Plant • Transformation cassettes are developed in the lab • They are then introduced into a plant • Two major delivery methods

  36. The Lab Steps

  37. Non-transgenics Transgenics The Next Test Is The Field Herbicide Resistance

  38. Before After Final Test of the Transgenic Consumer Acceptance RoundUp Ready Corn