Unit 6.4- Cloning and Biotechnology Flashcards
Clones definition:
Genetically identical organisms or cells
Vegetative propagation definition:
Natural plant cloning
Advantages of natural cloning:
- If the conditions of growth were good for the parent, they will be good for the offspring
- Rapid, the population can rise quickly to take advantage of suitable conditions
- Reproduction can be carried out even if there is only one parent and sexual reproduction is not possible
Disadvantages of natural cloning:
- Offspring may become overcrowded
- No genetic diversity apart from that caused by mutations
- Population shows little variation
- Selection is not possible
- If the environment is less advantageous, the whole population is susceptable
Runners or stolens:
- Horizontal stems from plants
- Can form roots at certain points
- Grow on the surface of the ground
Rhizomes:
- Horizontal stems from plants
- Can form roots at certain points
- Grow underground
- Some can be adapted into thicker overwintering organs from which one or more new stems will grow in the spring
Suckers:
- New stems that grow from the roots of a plant
- The horizontal branch may die, leaving the new stem as a separate individual
Bulbs:
- Overwintering
- Consist of an underground stem from which grow a series of fleshy bases
- There is an apical bud which will grow into a new plant in the spring
- There may be more than one apical bud, these each grow into separate new plants
Corms:
- Solid rather than fleshy
- Underground stem with scaly leaves and buds
- Remain in the ground over winter
- In the spring, the buds grow to produce one or more new plants
Leaves:
- Clones grow on the leaf margins
- The immature plants drop off the leaf and take root
Tubers:
Type of underground stem
Micropropagation definition:
Growing large numbers of new plants from a meristem tissue taken from a sample plant
Tissue culture definition:
Growing new tissues, organs or plants from certain tissues, organs or plants from certain tissues cut from a sample plant
How to clone plants by cuttings:
- Stem is cut between two nodes
- Cut ends placed in moist soil
- New roots will grow
- Some plants will need to be dipped in rooting hormone to help stimulate root growth
How to clone plants by micropropagation:
- Plant material cut into small pieces (explants)
- Explants are sterilised using dilute bleach or alcohol
- Explants placed on agar, containing nutrients and hormones such as auxin
- This stimulates the cells to divide by mitosis and for callus (mass of totipotent cells)
- Once a callus is formed, it is divided to form a large number of small clumps of undifferentiated cells
- Once plantlets have formed, these are transformed to a greenhouse to be grown in compost or soil and are acclimatised to normal growth conditions
Advantages of artificial cloning:
- Rapid
- Can be carried out when sexual reproduction is not possible
- Genetically identical to parent plant so posses same beneficial characteristics
- Uniform phenotype making them easier to grow and harvest
- Using the apical bud ensures plants are free from viruses
Disadvantages of artificial cloning:
- Tissue culture is labour intensive
- Expensive
- Can fail due to micro-bacterial contamination
- Cloned offspring are susceptible to the same pests and diseases. There can be a rapid spread of pests and diseases
- No genetic variation apart from that caused by mutations
Embryo twinning definition:
Splitting an embryo to create two genetically identical embryos
Enucleation definition:
Removal of the cell nucleus
Somatic cell nuclear transfer (SCNT) definition:
A technique that involves transferring the nucleus from a somatic cell to an egg cell
How does embryo splitting work?
- A zygote is created by IVF
- Zygote is allowed to divide by mitosis to form a small ball of cells
- The cells are separated and allowed to continue dividing
- Each small mass of cells is placed int the uterus of a surrogate mother
How does somatic cell nuclear transfer (SCNT) work?
- Enucleation of egg cell
- Somatic cell removed from animal to be cloned. Nucleus is removed
- Somatic cell nucleus fused with empty egg cell by applying electric shock
- Shock triggers egg cell to start developing as though it had just been fertilised
- Cell undergoes mitosis to produce small ball of cells
- Embryo placed in the uterus of a surrogate mother
What can therapeautic cloning be used for?
- Skin grafts
- Repair damage to spinal chord
- Restore capability to produce insulin in the pancreas
- Potential to grow whole new organs to replace diseased organs
Arguments for artificial cloning in animals:
- Can produce whole heard of animals with desirable characteristics
- Using genetically identical embryos for scientific research allows the effects of genes and hormones to be assessed with no interference from different genotypes
- Testing drugs on cloned cells and tissues avoids using people or animals for testing
- Can produce cells genetically identical to the donor, for use in repairing damage
- Individuals from endangered species can be cloned to increase numbers
Arguments against artificial cloning in animals:
- Lack of genetic variation can expose heard to diseases or pests
- Animals may be produced with little regard for their welfare
- Success rate of adult cell cloning is very low
- Expensive
- Ethical issues surrounding how long the embryo survives
- Cloning animals of endangered species may increase numbers, but doesn’t increase biodiversity
Biotechnology definition:
The use of living organisms or parts of living organisms in industrial processes.
Examples of food biotechnology is used in:
- Ethanol in beer and wine (yeast)
- CO2 used to make bread rise (yeast)
- Lactic acid used to make yogurt and cheese (bacteria)
- Mycoprotein (fungus)
- Soya beans are fermented to make soy sauce (yeast)
Examples of pharmaceutical drugs biotechnology is used in:
- Penicillin (fungus)
- Insulin (genetically modified bacteria)
Examples of enzymes biotechnology is used in:
- Biological washing powders (bacteria)
- Sucrase used to digest sugar to make food sweeter (yeast)
Advantages of using microorganisms in biotechnology:
- Cheap and easy to grow
- Lower temperatures and pressures
- Not dependant on climate so could happen anywhere in the world
- Short life cycle and reproduce quickly so large population can grow quickly
- Can be genetically modified easily
- Few ethical considerations
How is yogurt made?
- Milk is fermented
- The bacteria convert lactose in to lactic acid
- The acidity denatures the milk protein, causing it to coagulate
How is cheese made?
- Bacteria produces lactic acid from lactose in the milk
- The milk is then mixed with rennet, which contains rennin
- Rennin coagulates the milk protein (casein) in the presence of calcium ions
- Kappa-casein, which keeps the casein in solution, is broken down, making the casein insoluble
How is bread made?
- Flour, water, salt and yeast are mixed and needed
- Proving/ fermenting at warm temperature. Yeast respire anaerobically, producing CO2 bubbles, causing the dough to rise
- Cooking
How is wine made?
- Grapes have yeast on their skin
- When the grapes are crushed, they use the sugar in the grapes to produce CO2 and alcohol
How is beer/ale made?
- Barley grains are beginning to germinate (malting)
- This converts stored starch to maltose which is respired by the yeast and produces CO2 and alcohol
Advantages of using microorganisms in food production:
- Fast production
- High protein content
- No animal welfare issues
- Production can easily be increased/decreased in response to demand
- Not much land is required
Disadvantages of using microorganisms in food production:
- Some people won’t want to eat fungal protein or food that has been grown on waste
- Protein has to be purified to ensure it is not contaminated
What does scaling up drug production require?
Fermenters because conditions can be controlled to ensure the best possible yield
Conditions controlled in a fermenter:
- Temperature
- Nutrients available (carbon, nitrogen, vitamins, minerals)
- PH
- Concentration of product, if it is allowed to build up it might affect the synthesis process
What must a fermenter be sterilised with?
Superheated steam
Continuous culture:
- Primary metabolites are continually released from cells and can be extracted from the fermenting broth
- Broth is topped up with nutrients
- Some of the broth is removed regularly to extract the product and remove cells from the broth, so the product doesn’t become too dense
- The microorganisms grow at a continuous rate
Batch culture:
- Products produced when organism is under stress
- Secondary metabolites, and produced during the stationary phase of growth
- Culture is set up with a limited amount of nutrients and allowed to ferment for a specific amount of time
Asepsis definition:
Ensuring sterile conditions are maintained
Why is asepsis important?
-Unwanted microorganisms would reduce the production because they would compete for nutrients and space
-Reduce the yield of useful products
-Spoil the product
They may also:
-Produce toxic chemicals
-Destroy the cultural microorganisms and their products
Production of penicillin:
-Secondary metabolite produced once the PENECILLIUM CHRYSOGENUM population has reached a certain size
-Batch culture
1- Fermenter run for 6-8 days, then the culture is filtered to remove the cells
2- Precipitated as crystals by the addition of potassium compounds. May be modified by the action of other microorganisms or by chemical means
3-The antibiotic is mixed with inert substances and prepared for administration as a syrup, tablet, or in a form suitable for injection
Production of insulin:
- Bacteria are genetically modified
- Gene for human insulin combined with a plasmid to act as a vector, so the gene could be inserted into a bacterium
- Allows vast quantities of insulin to be produced at a relatively low cost
- Continuous culture
Bioremediation definition:
The use of organisms to clean the soil and underground water on a polluted site. The organisms convert the pollutants into less harmful substances
Conditions required for bioremediation:
- Available water
- Suitable temperature
- Suitable PH
- When conditions may not be suitable, the may be modified by the addition of suitable substances
- When conditions can not be made in situ, the soil may be dug up and treated ex situ
Advantages of bioremediation:
- Uses natural systems
- Less labour equipment needed
- Treatment in situ
- Few waste products
- Less risk of exposure to clean-up personnel
Disadvantages of bioremediation:
Only suitable for some products
Aseptic techniques:
- Washing hands
- Disinfect working area
- Have a bunsen burner operating nearby to heat air. Causes air to rise and prevents air-bourne microorganisms from settling
- As bottles are opened and closed, the neck should be flamed
- Don’t lift the lid of the petri dish off completely
- Any glass or metal equipment should be passed through a flame before it is used
What are the three main steps in growing microorganisms on an agar plate?
- Sterilisation
- Inoculation
- Incubation
How is agar sterilised?
It is heated in an autoclave at 121 C for 15 minutes. This kills all living organisms. When it has cooled enough to handle, it is poured into sterile petri dishes and left to set with the lid on
Inoculation definition:
The introduction of microorganisms to the sterile medium
What are the different methods of inoculation?
- Streaking: Wire loop used to transger a drop of liquid and then draw out by the wire loop
- Seeding: Pipette drops liquid before or after agar is added to the plate
- Spreading: Sterile glass spreader
- A small cotton swab can be moistened with distilled water and used to collect microorganisms from a surface and then wipe them over the agar
Closed culture definition:
A culture which has no exchange of nutrients or gases with the external environment
What are the stages of the growth curve:
- Lag phase
- Log phase
- Stationary phase
- Death phase
What happens in lag phase?
Small growth of microorganisms as there is a small population and they are still adjusting to their new environment. This may involve:
- Taking up water
- Cell growth
- Switching on certain genes
- Synthesising specific proteins
What happens in the log phase:
Quick growth because they have the enzymes needed to survive
What happens in the stationary phase?
- Nutrients used up
- Increasing amounts of waste products
- Population growth declines and death rate increases
- Eventually reproduction rate will equal death rate and there will be no population growth
What happens in death phase?
Nutrients run out and concentration of waste products may become lethal
Primary metabolites definition:
Produced during normal activities of the microorganism and collected during the log phase
Secondary metabolites definition:
Produced in the stationary phase. Collected at the end of the stationary phase or during the death phase
Immobilised enzymes definition:
Enzymes that are held in place and not free to diffuse through solution
Methods used to immobilise enzymes:
- Adsorption
- Covalent bonding
- Entrapment
- Membrane separation
How does adsoption work?
- Enzymes bound to supporting surface such as clay through hydrophobic interactions and ionic links
- The active site may be slightly distorted by the additional interactions affecting enzyme activity
How does covalent bonding work for immobilising enzymes?
- Enzyme molecules are bonded to supporting surfaces such as clay through covalent bonds
- They are bonded using a cross linking agent which may also link them in a chain
- Can be expensive and may distort active site, reducing activity
- Enzymes are much less likely to become detached and leak into the reaction mixture
How does entrapment work?
- Molecules are trapped in a matrix that does not allow free movement
- The molecule is unaffected and remains fully active
- Only suitable for processes where the substrate and product molecules are relatively small so they can diffuse in and out of the matrix
How does membrane separation work?
- Enzyme molecules separated from reaction mixture by a partially permeable membrane
- The substrate and product molecules must be small enough to pass through the membrane by diffusion
Industrial uses of immobilised enzymes:
- Glucose isomerase
- Penicillin acyclase
- Lactase
- Aminoacylase
- Glucoamylase
- Nitrile hydratase
How are immobilised enzymes used with penicillin acyclase?
Formation of semi-synthetic penicillins. Some penicillin resistant microorganisms are not resistant to these semi-synthetic forms
How are immobilised enzymes used with lactase?
- Convert lactose to glucose and galactose by hydrolysis
- Used to produce lactose free milk
