Plant Biotech

• Plant Biotechnology


  The application of laboratory-based techniques for plant propagation or genetic improvement
• Plant tissue culture

  • Micropropagation
  • Biotech for producing plants
  • Suspension cultures
  • Production of valuable compounds
  • Somatic hybridization
  • Production of novel germplasm
• Micropropagation


  1. Starts with an explant which is cultured under sterile conditions designed to promote regeneration of whole plants

  2. The ability to regenerate whole plants from explants depends on the totipotency of many somatic plant cells

  3. Totipotency: the ability of an individual cell form all parts of the mature organism

  4. Depends on plant cells' capacity for dedifferentiation and redifferentiation
• Regeneration by organogenesis


  1. Isolation of an explant under sterile conditions

  2. Callus production on nutrient medium containing plant hormones (auxin and cytokinin – root and shoot growth)

  3. Organogenesis stage I: the generation of new shoots from the undifferentiated callus (promoted by cytokinin)

  4. Organogenesis stage II: the generation of roots from the shoots (promoted by auxin)
• Regeneration by somatic embryogenesis


  1. Convince callus to become embryo-like – known as somatic embryos

  2. Somatic embryogenesis = development of embryos and whole plants directly from somatic cells

  3. Embryogenic callus -> somatic embryo -> plantlet
• Somaclonal variation

  • Phenotypic variability between individual plants derived from plant tissue culture
  • Genetic changes: polyploidy and aneuploidy, chromosome structure and DNA sequence
  • Advantages: creation of additional genetic variability for plant improvement
  • Disadvantages: lack of uniformity is a problem in micropropagation for horticulture and forestry industries
• Cell suspension cultures


  • The growth of plant cells under sterile conditions in a liquid medium, with shaking
  • Used for research: convenient method for obtaining a homogeneous mass of cells in the lab
  • Used for commercial production of high-value secondary metabolites and other compounds e.g. antimicrobials, vitamins and food flavours
• Somatic hybridization using protoplasts

  Somatic hybridization: production of novel hybrids between sexually incompatible plant species

  1. Protoplasts from the two species are mixed and fused to produce hybrid protoplasts

  2. Plants are regenerated from the hybrid protoplasts by tissue culture

  3. Allow plants that you wouldn't be able to cross sexually

  4. Example Shanrong No.3 (SR3) cross between common wheat and tall wheatgrass

  5. Plants are sterile and you can't propagate them
• Producing a genetically engineered or 'transgenic' plant

  1. Production of a DNA construct that harbours the gene of interest

  2. Transformation of plant cells with the construct

  3. Selection of transformed cells using a selectable marker

  4. Regeneration of whole plants from the transformed cells using organogenesis or somatic embryogenesis
• Promoter

  • Constitutive (on all the time throughout the plant)
  • Tissue-specific (e.g. only expressed in root hairs)
  • Developmentally regulated (e.g. activated during fruit ripening)
  • Inducible (only activated when a chemical treatment is applied)
• Selectable marker


  Genes that confer resistance to compounds that are toxic to plants, e.g. Antibiotic-resistance, Herbicide-resistance
• Plant transformation: delivery of DNA into plant cells

  1. Naked DNA delivery systems, e.g. Particle bombardment, Electroporation

  2. Natural delivery systems, e.g. Agrobacterium tumefaciens
• Naked DNA delivery by particle bombardment

  Coat DNA with gold, Gold is sent into the particles. Can generate new DNA. Gene gun uses helium
• Naked DNA delivery by electroporation

  1. Use of a short, high voltage electrical discharge to make the plasma membrane permeable to DNA (or other polar molecules), membrane reseals quickly

  2. Usually used with plant protoplasts
• Agrobacterium tumefaciens


  • Plant pathogen that infects wound sites and produces tumours (crown gall disease) in many plant species
  • Able to transfer a small segment of its DNA (the 'T-DNA') into the plant genome
  • The T-DNA carries genes that induce uncontrolled cell division ('tumorigenic') and direct the synthesis of opines - amino acids the bacterium (but not the plant) can use as nutrients
• Ti plasmid

  • The T-DNA is bounded by two regions called the left and right borders (TL and TR )
  • Both borders are flanked by a 25-bp direct repeat sequence
  • The borders + repeats are the only sequences necessary for integration into the plant DNA
  • Vir gene are used for infection. And recognise left and right borders
• Floral dipping


  A method of plant transformation where developing flowers are immersed in a suspension of Agrobacterium – requires no tissue culture
• Transgenic crops

  • Modify the expression of existing genes (over-expression, reduced expression / stop expression, altered location/timing of expression)
  • Add new genes from other plants or other organisms
  • Delete genes – gene editing
• Targets of plant biotech

  • Agronomic Traits (Yield, Herbicide tolerance, Pest and disease resistance, Abiotic stress resistance, Reproduction)
  • Novel Crop Products (Oils, Proteins, Polymers)
  • Quality Traits (Processing, Shelf-life, Nutritional quality, Reduced anti-nutritionals)
  • Biofuels
• Herbicide tolerance - roundup-ready


  • Glyphosate (Roundup®) is a broad-spectrum herbicide made by Monsanto since the 1970s
  • Acts by inhibiting a chloroplast enzyme (EPSP synthase) required for aromatic amino acid biosynthesis
  • A mutant EPSP synthase gene (shkG*) was isolated from a bacterium resistant to the herbicide, and introduced into plants to confer glyphosate resistance
• Pros and cons of herbicide tolerance crops

  • Pros: Enhanced crop productivity, improves soil health, encourages 'no-till' agriculture
  • Cons: Increased use of RoundupTM, possible reduction in biodiversity
• Bacillus thuringiensis -toxin


  • A soil bacterium whose spores contain a crystalline protein (Bt toxin, encoded by cry genes) that kills insects
  • Very specific – narrow target range: the toxin binds to specific receptors in the gut of the target insect - so not toxic to other animals
• How bacillus thuringiensis works

  1. Larva consumes toxin

  2. Crystals solubilized and toxin activated by proteolytic cleavage in mid-gut

  3. Toxin binds to specific receptors

  4. Binding triggers cell death in mid-gut lining

  5. Septicaemia follows and larva dies
• Engineering plants with the BT gene

  • Different cry genes target different groups of insects
  • DNA constructs carrying different cry genes have been introduced into maize, cotton and other crops
• Pros and cons of Bt crops

  • Pros: Reduced financial costs, reduced environmental impact, improved health of farmers, improved food quality
  • Cons: Selection pressure for resistant pests, possible effect on non-target insects
• Vitamin A

  • Retinol is a pigment of eye, growth hormone
  • Humans obtain their vitamin A from provitamin A (β-carotene) from plants, and from animal products containing vitamin A
• Vitamin A deficiency (VAD)

  • Symptoms: night-blindness, xerophthalmia ('dry eyes')
  • Associated with increased susceptibility to death
  • A severe public health problem in many developing countries
• Making rice rich in beta carotene

  1. Introduced two daffodil genes and an Erwinia gene (from bacteria) under the control of endosperm-specific promoters into rice

  2. The resultant transgenic rice lines had enhanced β-carotene levels in the endosperm – 'Golden Rice'
• Golden rice

  • GR2 rice contains sufficient β-carotene to provide 50% of a child's RDA of vitamin A
  • Incorporated into rice breeding programmes in Philippines, India, Bangladesh, China and Vietnam
• Pros and cons of golden rice

  • Pros: The potential to save millions of children from death or blindness
  • Cons: Reinforces dependence on rice diet, better to increase diversity in agriculture and diet
• Purple fruit

  • Anthocyanin provides colour of red cabbage, cranberries and blueberries
  • Anthocyanins are good antioxidants and reduce the incidence of cancer and obesity in mouse disease models
  • Natural production in oranges only happens at a narrow temperature range – limited production is possible
• Non-browning fruit and veg

  • Arctic apples and Innate potatoes both have reduced polyphenol oxidase, reducing bruising
  • Innate potatoes also have reduced levels of Asparagine, which at high temperatures can be converted to acrylamide, a carcinogen
• CRISPR (Cas9)

  • Bacterial defence against viruses
  • Bacteria use short guide RNA (CRISPR) plus enzyme (cas9) to cut viral DNA
  • Introduce artificial guide RNAs to target genes of interest
  • In the host cell random repair errors introduce mutations
  • Can engineer specific sequence changes, insertions or deletions
• use of micropropagation


  propagating plant genotypes where other propgation methods are too difficult
• What is organogenesis?

  sequential culture to become a whole plant
• Describe a terminator sequence

  Made up of polyadenylation sequences
• What does E.coil nptII gene code for

  neomycin phosphotransferase which inactivates antibiotics such as kanamycin and neomycin
• When is Naked DNA transfer suitable?

  both dicots and monocots
• How to carry out a plant transformation using agrobacterium tumefaciens

  T-DNA containing foreign gene invades leaf which is transferred to agar dish containing hormone and antibiotic

  planft containing Ti plamsid will grow
• Describe Bt maize vairieties (MON810)


  Cry1Ab gene confers resistance against european stem borer