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    The science-of-transgenics The science-of-transgenics Presentation Transcript

    • The Science of Transgenics Phil McClean Department of Plant Science North Dakota State University The Sociological, Economic, and Ethical Impact of Transgenic Organisms Workshop Fargo, ND February 21, 2003
    • 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
    • What About the Term Genetic Engineering? Genetic engineering involves:
      • Isolating genes
      • Modifying genes so they function better
      • Preparing genes to be inserted into a new species
      • Developing transgenes
      Genetic engineering is the basic tool set of biotechnology
    • What is a transgenic? 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 Concept Based on the Term Transgene
    • We can develop organisms that express a “novel” trait not normally found in the species Why are transgenics important? Extended shelf-life tomato (Flavr-Savr) Herbicide resistant soybean (Roundup Ready)
    • Agriculture Transgenics On the Market Source: USDA
      • Insect resistant cotton – Bt toxin kills the
      • cotton boll worm
      • transgene = Bt protein
      • Insect resistant corn – Bt toxin kills the
      • European corn borer
      • transgene = Bt protein
      Normal Transgenic
      • Virus resistance - papya resistant to
      • papaya ringspot virus
      • transgene = virus coat protein
      Source: Monsanto
      • Herbicide resistant crops
      • Now: soybean, corn, canola
      • Coming: sugarbeet, lettuce, strawberry
      • alfalfa, potato, wheat (2005?)
      • transgene = modified EPSP synthase or
      • phosphinothricin-N-acetyltransferase
      • Biotech chymosin; the enzyme used
      • to curdle milk products
      • transgene = genetically engineered enzyme
      • bST; bovin somatotropin; used to increase
      • milk production
      • transgene = genetically engineered enzyme
      Source: Rent Mother Nature Source: Chr. Hansen
    • Some Ag Biotech Products Are Discontinued
      • Poor Quality
      • FlavrSavr tomatoes (Calgene)
      • Negative Consumer Response
      • Tomato paste (Zeneca)
      • Negative Corporate Response
      • NewLeaf (Monsanto)
      • Universal Negative Publicity
      • StarLink corn (Aventis)
    • Next Generation of Ag Biotech Products Source: Minnesota Microscopy Society Golden Rice – increased Vitamin A content (but not without controversy) transgene = three pathway enzymes Sunflower – white mold resistance transgene = oxalate oxidase from wheat
    • Turfgrass – herbicide resistance; slower growing (= reduced mowing) Bio Steel – spider silk expressed in goats; used to make soft-body bullet proof vests (Nexia)
    • Biotechnology is Not Just on the Farm Disease Treatment Diagnostics   Environmental Cleanup   Human Applications
    • 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
    • Biotechnology and Health The genes for these proteins are:
      • Cloned
      • Inserted into bacteria
      • Product isolated using biofermentation
      Product Use Insulin Diabetes Interferon Cancer Interleukin Cancer Human growth hormone Dwarfism Neuroactive proteins Pain
    • Environmental Applications Bioremediation - cleanup contaminated sites; uses microbes designed to degrade the pollution Indicator bacteria – contamination can be detected in the environment
    • Tooth decay – engineered Streptococcus mutans, the bacteria that destroys enamel Future Health-related Biotech Products Vaccines – herpes, hepatitis C, AIDS, malaria
    • 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
    • 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
      • Some claim any improved biological product is a GMO
      • They feel this will
      • For example, some call plant varieties biotechnology products
      • This is a false claim
      The GMO Ruse
      • ease the publics fear
      • pave the way for product acceptance
    • 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
    • Important Plant Improvement Methods
      • Breeding
      • Crossing two individuals from the same species;
      • produces a new, improved variety;
      • not a biotechnology procedure
      • Transformation
      • Adding a gene from another species; the
      • essential biotechnology procedure to produce
      • transgenics
      Source: USDA Source: USDA
    • Wheat Rye Triticale X Interspecific Cross New species, but NOT biotechnology products
    • 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
    • ATTCGA ATT G GA Susceptible Normal Gene Resistant Mutant Gene Mutagenesis Treatment Mutagenesis Changes the DNA Sequence
    • 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
    • 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
    • + Glyphosate X Roundup Sensitive Plants X X Without amino acids, plant dies X Shikimic acid + Phosphoenol pyruvate 3-Enolpyruvyl shikimic acid-5-phosphate (EPSP) Plant EPSP synthase Aromatic amino acids
    • Bacterial EPSP synthase Shikimic acid + Phosphoenol pyruvate 3-enolpyruvyl shikimic acid-5-phosphate (EPSP) Aromatic amino acids Roundup Resistant Plants + Glyphosate With amino acids, plant lives RoundUp has no effect; enzyme is resistant to herbicide
    • 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
    •  -Carotene Pathway Problem in Plants IPP Geranylgeranyl diphosphate Phytoene Lycopene  -carotene (vitamin A precursor) Phytoene synthase Phytoene desaturase Lycopene-beta-cyclase ξ-carotene desaturase Problem: Rice lacks these enzymes Normal Vitamin A “ Deficient” Rice
    • The Golden Rice Solution Daffodil gene Single bacterial gene; performs both functions Daffodil gene  -Carotene Pathway Genes Added IPP Geranylgeranyl diphosphate Phytoene Lycopene  -carotene (vitamin A precursor) Phytoene synthase Phytoene desaturase Lycopene-beta-cyclase ξ-carotene desaturase Vitamin A Pathway is complete and functional Golden Rice
    • Introducing the Gene or Developing Transgenics Steps 1. Create transformation cassette 2. Introduce and select for transformants
    • Transformation Cassettes Contains 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 Steps Prepare tissue for transformation Introduce DNA Culture plant tissue Field test the plants
      • Agrobacterium or gene gun
      • Multiple sites, multiple years
      • Develop shoots
      • Root the shoots
      • Leaf, germinating seed, immature embryos
      • Tissue must be capable of developing into normal plants
      • Transformation cassettes are developed in the lab
      • They are then introduced into a plant
      • Two major delivery methods
      Delivering the Gene to the Plant
      • Agrobacterium
      • Gene Gun
      Tissue culture required to generate transgenic plants
    • The Lab Steps
    • The Next Test Is The Field Herbicide Resistance Non-transgenics Transgenics
    • Final Test of the Transgenic Consumer Acceptance RoundUp Ready Corn Before After