6.4 Cloning and biotechnology

• Natural clones in plants

  • Genetically identical offspring produced by asexual reproduction. usually occurs by vegetative propagation (different parts of the plant grow into new individuals)
• Vegetative Propagation

  1. Runners - stems grow horizontally, form when they touch the soil (strawberry)
  2. Rhizomes - Underground horizontal stems, develop into new shoots and roots (Ginger, grass)
  3. Tubers - swollen underground stems (potatoes)
  4. Bulbs - Underground storage organs (onions)
  5. Corms - solid underground stem bases, can produce new shoots each seasons
  6. Suckers - new shoots that grow from the roots of parent plant
  7. Leaves - drop off and root
• How to take plant cuttings

  1. Use a healthy shoot
  2. Stem is cut at a slant between leaf and the nodes
  3. dip in rooting powder
  4. Place in soil and add water
  5. to reduce transpiration, cover with plastic bag
• Production of artificial clones by micropropagation

  1. Small tissue samples (explants) are taken from parent plant, typically from meristem cells; totipotent and can differentiate into any cell
  2. Sterilisation to remove and inhibit growth of bacteria and fungi
  3. Cultured on a nutrient rich medium
  4. divide to form undifferentiated mass of cells called a callus
  5. Callus transfered to a new medium with conditions to encourage shoot and root formation - allows them to differentiate into plantlets
  6. Plantlets moved to soil to develop into mature identical plants
• Evaluation of micropropagation

  Pros -

  • rapid and large scale
  • disease free clones
  • genetically identical
  • space efficient

  Cons-

  • Genetically identical - vulnerable to disease and environmental changes
  • Expensive and needs skilled technicians
• Natural clones in animals

  1. Monozygotic Twins - fertilised egg splits into two genetically identical embryos. grow independently
  2. Invertebrates - Regeneration/fragmentation; forms genetically identical offspring from parts of body that have broken off
• Artificial embryo twinning

  Single embryo manually splits before cells start to differentiate

  1. Female is treated with hormones to produce multiple eggs
  2. Ova are extracted and fertilised in Petri dish - embryo
  3. Divides into several cells, still totipotent
  4. Each cell in its own dish and develop into individual embryos
  5. Implanted into uteruses
• Enucleation

  Removing the nucleus from a cell
• Somatic cell Nuclear Transfer

  1. Somatic cell taken from the organism
  2. unfertilised egg taken from another female and its nucleus is removed by enucleation
  3. Nucleas of the somatic cell inserted into the enucleated egg cell which now contains diploid DNA of the donor
  4. Egg is shocked with electricity tigger mitosis
  5. Grows into an embryo - until the blastocyst stage then implanted in mother
• Uses of animal cloning

  1. Drug testing and disease modelling as there's no genetic variation to impact results
  2. Conservation - boost the numbers from a limited gene pool
  3. Agriculture - replicate animals with desirable characteristics
  4. Provide source of immunocompatible stem cells
• For and against animal cloning

  For -

  • Transmission of desirable characterises
  • Enable reproduction if infertile
  • preserve biodiveristy
  • rapidly increase size of population

  Against -

  • high cost and complex
  • reduces genetic diversity
  • ethical concern destroying embroys
  • SCNT inefficient - may take many ova to produce single clone; high failure rate
• Use of microorganisms in biotechnology (waste treatment)

  Use microbes to clean up pollutants

  • Oil spills; pseudomonas break down hydrocarbons
  • Sewage
• Use of microorganisms in biotechnology; For and Against

  1. Low Costs; require low temperature or few energy requirements. nutrients for growth are cheap
  2. Large numbers can be produced; reproduce quickly
  3. better for the environment as there's less pollution; reduces use of land for food production and has lower energy requirements
  4. Can be produced in many locations; not affected by climate
  5. More healthy food as its low in cholesterol and high in protein. easy to genetically engineer
• How to culture microorganisms using aseptic techniques

  1. Sterilisation - Medium heated in an autoclave at 121 degrees fro 15 mins - kills all living organisms. poured into Petri dish
  2. Have a bunsen buner - heats the air, air rises and prevents airborne microorganismss
  3. Inoculation - introduction of microorganisms.
  4. All equipment sterilised by heating
  5. Incubation - Petri dish labelled and tapes. not sealed completely otherwise anaerobic bacteria (may be pathogenic) will be introduced. placed upside, prevent condensation falling onto agar
• Agar

  Polysaccharide of galactose from seaweed to thicken medium into gel
• Batch vs Continuous Fermentation

  .

  A) closed

  B) competition

  C) stationary

  D) easy

  E) less

  F) removed

  G) log

  H) difficult

  I) more

  
• Standard growth curve in a closed culture

  .

  A) lag

  B) log

  C) stationary

  D) death

  
• Growth pattern phases

  1. Lag - slow initial cell growth as they adapt to environment
  2. Log - rapid doubling of cell under ideal conditions, growth rate at maximum
  3. Stationary phase - plateaus as nutrients diminish and waste accumulates
  4. Death - cell death rate exceed cell growth rate due to limited resources and increase in toxins
• Factors affecting growth of microorganism

  1. Temp
  2. pH
  3. Nutrient availability
  4. Antimicrobial substances
• Immobilised enzymes

  Enzyme attached to an inert material in order to restrict movement, not free to diffuse
• Main Methode of enzyme immobilisation

  1. Binding - covalent bonds using cross-linking agent. can be expensive and may distort enzyme active site
  2. Adsorption - enzymes may be adorned onto the surface on an insoluble support by hydrophobic interactions and ionic links
  3. Entrapment - trapped in a matrix, remain fully active but substates and products must diffuse in and out matrix. only suitable for molecules that are small
• Why are immobilised enzymes more stable over a range of pH than free ones

  Less likely to denature as they aren't moving freely. Active site remains in tact and fully functional
• Adv and Disadv of immobilised enzymes

  Advantages -

  • Cost effective - can be reused, no need to purchase more
  • Produce enzyme free products
  • more tolerant of temp and pH changes

  Disadvantages -

  • High initial costs
  • may reduce enzyme activity
  • reactor systems are complex and prone to technical problems