Cloning and biotechnology Flashcards
Clones
-Genetically identical copies of organisms/cells
-Produced asexually in which the nucleus is divided by mitosis
How are clones produced by mitosis
-Mitosis creates two genetically identical copies of the DNA
-The DNA is separated into two genetically identical nuclei
-Before the cell divides and creates two genetically identical cells
Why may the cells produced by mitosis not be identical
May not be physically/chemically identical as after division they may differentiate to form two different types of cell
How does yeast/ bacteria reproduce
Asexually by budding
Asexually by binary fission
Advantages of natural cloning
-If conditions of growth is good for the parent it will be good for the offspring
-Cloning is rapid so population can reproduce quickly to take advantage of the suitable environment conditions
-Reproduction can be carried out when sexual reproduction may not be possible
Disadvantages of natural cloning
-Offspring may become over crowded
-No genetic diversity ( except from mutations in the DNA)
-Population shows little variation
-Selection is not possible
-If environment changes to be less advantageous, the whole population is susceptible
Vegetative propagation
Natural cloning
Reproduction from vegetative parts of the plant - usually an over wintering organ rather than through specialised reproductive structures
Plant cloning
Differentiation of many plant cells is not as complete as in animals many parts of the plant contain cells with the ability to divide and differentiate into other types of cell
Examples of plant cloning by vegetative propagation
-Runner/ stolens/ rhizomes/ suckers
-Bulbs
-Corms
-Leaves
-Runners/ stolens / Rhizomes
Plants that grow horizontal stems that can grow roots at any point
-Grow on the surface of the ground (Runners/ stolens)
-Grow under ground (Rhizomes )
How are some Rhizomes adapted
Adapted as thickened overwintering organs from which one or more new stems will grow in the spring
Suckers
New stems that grow from the roots of the pant - may be close to the base of the older stem/ some distance away
-Original horizontal branch will die and leave the new stem as a separate individual
Bulbs
e.g. onions
-Overwintering mechanism
-Consists of an underground stem which grow a series of fleshy leaf bases
- Also an apical bud that will grow into a new plant in the spring
-Often bulb contains more than one apical bud and each will grow into a new plant
Corms
-Solid rather than fleshly like a bulb
-Underground stem with scaly leaves and buds
-Remain in the ground over winter
-In spring the buds grow to produce one or more plants
Leaves
Kalanchoe plant reproduces asexually as clones grow on the leaf margins
-Immature plants drop off the leaf and take root
Tubers
Another type of underground stem i.e. potatoes
-One potato will grow into one/more plants
-Each new plant can produce many new tubers (potatoes) later that year
Cloning in animals
Don’t clone as often as plants but there are some e.g. of natural cloning i.e. identical twins
How do identical twins form
Fertilised egg (zygote) divides as normal but the two daughter cells split to become two separate cells
-Each cell grows and develops into a new individual
What is the easiest way gardeners/growers make clones
Cuttings
-A stem is cut between the nodes
-Cut end of the stem is placed in moist soil
-New roots will grow from the tissue in the stem - usually from the node but may grow from other parts of the buried stem
What helps stimulate root growth from cuttings
Dipping the cut stem in rooting hormone / remove the bark from the cut end of the stem as this encourages the plant to produce a callus
Where else can a cutting be taken from a plant
-Root cuttings = section of the root is buried just below the soil surface and produces new shoots
-Scion cuttings = dormant woody twigs
-Leaf cuttings = leaf placed on moist soil ; leaves develop new stems and roots
What are the disadvantages of taking cuttings for natural cloning
Large scale cloning can be time time-consuming and take a lot of space
-Some plants don’t respond well to cuttings
Tissue culture
Growing new tissues, organs, plants from certain tissues cut from the sample plant
-Alternative to taking cuttings
-Carried out on a nutrient medium under sterile conditions
-Application of plant growth substances at the correct time can encourage the cells in the growing tissues to differentiate
What is tissue culture most used for
commercially used to increase the number of new plants in micropropagation
Micropropagation
Taking a small piece of plant tissue (meristem) and using plant growth substances to encourage it to grow and develop into a new plant
What are the six steps that micropropagation involves
(First step)
1) Suitable plant material is selected and cut into small pieces = explants
-Could be tiny pieces of leaf/stem/bud
-Meristem tissue used as free from virus infection
What are the six steps that micropropagation involves
(Second step)
2) Explants are sterilised using dilute bleach/alcohol
-Kills any bacteria/ fungi as these would thrive in the conditions required for the plant to grow well
What are the six steps that micropropagation involves
(Third step)
3) Explants placed on a sterile growth medium (usually agar gel) containing suitable nutrients i.e. glucose/ amino acids/ phosphates
-Gel also contains high concentrations of the plant growth substances auxin/cytokines
-Stimulates the cell of each explant to divide by mitosis to form a callus
What is a callus
A mass of undifferentiated totipotent cells
What are the six steps that micropropagation involves
(fourth step)
4) Once callus has formed it is divided to produce a larger number of small clumps of undifferentiated cells
What are the six steps that micropropagation involves
(fifth step)
5) Small clumps of cells are stimulated to grow, divide and differentiate into different plant tissues
=Achieved by moving cells to a diff growth media
Why do you need to move the cells to a diff growth media
(fifth step)
As each media contains diff ratio of auxin and cytokines
-First 100 auxin:1 cytokine stimulates roots to form
-Second 4 auxin: 1 cytokinin stimulates shoots to form
What are the six steps that micropropagation involves
(sixth step)
6) Once tiny plantlets have formed they are transferred to a greenhouse to be grown in compost/soil and acclimatised to normal growing conditions
Seed banks (tissue culture)
Use tissue culture techniques to store plants at a growth stage when they are not too large
-Technique important in the conservation of species whose seeds do not remain viable for long periods
Advantages of artificial cloning (1-3)
1) Rapid method of growing new plants rather than growing plants from a seed
2) Cloning can be carried out when sexual reproduction is not possible.
-Plants that have lost their ability to breed sexually can be re-produced i.e. commercially grown bananas
-Plants that are hard to be grown from seed can be reproduced i.e. orchids
3) Plants selected will all be genetically identical to the parent plant = display the same desirable characteristics such as high yield / disease resistance/ colour
Advantages of artificial cloning (4-6)
4) If original plant had unusual characteristics due to selective breeding / genetic modification then combination will be retained without the risk of losing it through sexual reproduction
5) New plants are all uniform in their phenotype = easier to grow and harvest
6) Using the apical bud (meristem) as an explant for tissue culture ensures the new plants are free from viruses
Disadvantages of artificial cloning (1-3)
1) Tissue culture labour is extensive
2) Expensive to set up the facilities to perform tissue culture successfully
3) Tissue culture could fail due to microbial contamination
In animals what does successful cloning begin with
Cells that are totipotent
Totipotent
Cells that can divide and differentiate into all types of cell found in an adult organism
-In animals theses are very early embryo cells
Reproductive cloning of animals may be useful for:
- Selective breeding/ genetic modification
-Genetically modified animals may develop unusual characteristics i.e. cows that produce less methane
Two main techniques to achieve reproductive cloning
-Embryo twinning: splitting an embryo to make two genetically identical embryos
- Somatic cell nuclear transfer (SCNT): A technique that involves transferring the nucleus from a somatic egg to an egg cell
Embryo splitting
1) A zygote is produced in vitro fertilisation from a male and female with desirable characterises
2) Zygote allowed to divide by mitosis to create a small ball of eggs
3) Cells are separated and allowed to continue to divide
4) Each small mass of cell is placed into the uterus of a surrogate mother
SCNT advantages
The only way to clone an adult
Advantage: phenotype known before the cloning starts
SCNT
1) Enucleation: egg cell obtained and nucleus is removed
2) Normal body cell (somatic cell) from the adult to be cloned is isolated and the nucleus is removed
3) Nucleus of adult cell fused with empty egg cell by an electric shock
4) Shock also triggers egg cell to start developing as if it has been fertilised
5) Cell undergoes mitosis to produce a small ball of cells
6) Young embryo placed in the uterus of a surrogate mother
Non-reproductive cloning
The production of cloned cells and tissues for purposes other than reproduction:
1) Therapeutic cloning
2) Cloning for scientific research
Therapeutic cloning
New tissues/ organs can be grown as replacements for people who are not well
Therapeutic cloning e.g.
-Skin can be grown in vitro to act as a graft for skin burns
-Cloned cells can repair damage to the spinal cord of a mouse and restore its capability to produce insulin in its pancreas
-Potential to grow whole new organs to replace diseased organs
What is the advantage of tissues grown from the patients own cells
Less chance of rejection from the body’s immune system
Cloning for scientific research
-Useful for research into the action of genes that control development and differentiation
-Also used to grow specific tissues/organs for use in tests on the effects of medicinal drugs
Arguments for artificial cloning in animals
1) Produces whole herd of animals with a high yield/ shows an unusual combination of characteristics i.e. silk in their milk (lambs)
2) Produces genetically identical individuals of high value characteristics
3) Identical embryos/ tissues are useful in assessing the effect of genes and hormones with no interference from different genotypes
4) Testing medicinal drugs on clones stops drugs being tested on humans/animals
5) Can produce cells genetically identical to the donor to repair tissue damage from accidents
6) Individuals from an endangered species can be cloned to increase numbers
Arguments against artificial cloning in animals
1) Lack of genetic variation may expose the herd to certain diseases/pests
2) Animals may be produced with little regard for their welfare, which may result in meat producing chickens that cannot walk (unethical)
3) Success rate of adult cloning is poor and its a lot more expensive then conventional breeding
Arguments against artificial cloning in animals
1) Lack of genetic variation may expose the herd to certain diseases/pests
2) Animals may be produced with little regard for their welfare, which may result in meat producing chickens that cannot walk (unethical)
3) Success rate of adult cloning is poor and its a lot more expensive then conventional breeding
4) Cloned animals may be less healthy and have shorter life spans
5) Ethical issues regarding how long the embryo survives and whether it is right to create a life and simply destroy it
6) If endangered species are cloned to increase numbers it does not help increase genetic diversity
Biotechnology
The use of living organisms/ parts of living organisms in industrial processes to produce food, drugs or other products
What are the four main areas, in which micro-organisms are used in biotechnology
1) Food
2) Pharmaceutical drugs
3) Enzymes
4) Other products
Microorganisms used in food production
-Ethanol in beer and wine (yeast)
-CO2 to make bread rise (yeast)
-Lactic acid to make bread/cheese (Lactobacillus)
Microorganisms used in pharmaceutical drugs
-Penicillin (penicillium fungus)
Microorganisms used in enzymes
-Protease/lipase in washing powder (bacteria)
Advantages of using micro-organisms in biotechnology
1) Cheap/ easy to grow
2) Production takes place at lower temperatures (saves on fuel and energy)
3) Production takes place at normal atmospheric pressure (safer than using chemical reactions which requires high pressure)
4) Production not dependent on climate (can take place anywhere with suitable resources and equipment)
5) Micro-organisms can be fed by products from other food industries i.e. starch, waste, water
6) Short life cycle/ reproduce quickly
What is the reaction vessel called
Fermenter: vessel used to grow populations of microorganisms
What are other organisms are used in biotechnology
Genetically modified animals to produce useful products
Other forms of biotechnology
-Gene technology
-Gene modification snd therapy
-Selective breeding
-Cloning by embryo splitting / micropropogation
-use of enzymes in industrial processes
-Immunology
What types of food are micro-organisms in
-Yogurt
-Cheese
-Baking
-Alcohol beverages
Yogurt what micro-organisms are involved
Milk that has undergone fermentation (lactobacillus bulgaricus) (Streptococcus thermophilus)
How is yogurt made
1) Bacteria convert lactose to lactic acid
2) Acidity denatures the milk protein causing it to coagulate
3) Bacteria partially digests the milk making it easy to digest
Fermentation also produces the flavour characteristics of yogurt
Coagulate
(of a fluid, especially blood) change to a solid or semi-solid state
Bacteria in yogurt as probiotics
L. acidophilus
-May benefit human health by improving digestion of lactose
-Aids gastrointestinal function and stimulating the digestive system
Cheese - how is it made (1)
1) Milk is pre-treated with a culture of bacteria (Lactobacillus) that can produce lactic acid from lactose
2) Once acidified the milk is mixed with rennet
3) Rennet contains enzyme Rennin which coagulates the milk protein (casein) in the presence of calcium ions
Cheese - how is it made (2)
What happens when the milk is mixed with Rennet?
1) Kappa- casein (keeps casein in solution) is broken down. Casein insoluble
2) Casein is precipitated by the action of Calcium ions, which binds the molecules together
Cheese - how is it made (3)
Curd
1) Resulting solid: curd is separated from the liquid component by stirring and heating
2) Bacteria continues to grow and produces more lactic acid
3) Curd is then pressed into moulds
Cheese - how is it made (4)
Treatment while pressing the curd
This determines the characteristics of the cheese
Flavour determined by the later ripening and maturing process
Cheese - how is it made (4)
How is extra flavour added
Inoculation with fungi (e.g. penicillium) to produce ‘blue’ cheese
Bread - what is it (1)
A mixture of flour; water; salt; yeast (Single celled fungus)
Bread - how is it made (2)
1) Mixing - ingredients are mixed thoroughly together by kneading
2) Proving/fermenting - Dough is left in a warm place for up tot three hours whilst the yeast respires anaerobically. Produces CO2 which causes the bread to rise
3) Cooking - the risen dough is baked. Any alcohol evaporates during the cooking process
Alcoholic beverages
The product of the anaerobic respiration of yeast (S .cerevisiae)
Wine - How is made
Made using grapes that have natural yeasts in skin
-Grapes contain sugars/ fructose/ glucose
When grapes are crushed the yest, uses these sugars to produce CO2 and alcohol
Ale/ beer - How is it made?
Brewed by barley grains that are beginning to germinate (malting)
-As grain germinates it converts starch to maltose, which is respired by the yeast
-Anaerobic respiration produces CO2 and alcohol
-Hops used to give a bitter taste to the liquid
Single celled protein (SCP)
Fungal protein / mycoprotein used in food
-Quorn
-Can produce protein with a similar amino acid profile to animal/plant protein
-Grow on almost any organic substrate, including waste materials such as paper and whey (curdled milk from which the curds have been removed)
Advantages of using micro-organisms in food (1)
1) Production of protein faster to produce than animal/plant protein
2) Biomass produced has a high protein content (48-85%)
3) Production can be increased/decreased accordingly
4) No animal welfare issues
5) Micro-organisms - good source of protein
Advantages of using micro-organisms in food (2)
6) Micro-organisms genetically modified to adjust the amino acid content of a protein
7) SCP production can be combined with the removal of waste products
8) Production independent of weather
9) Not much land required
Disadvantages of using micro-organisms in food (1)
1) Some people may want to eat fungal food/ food grown on waste
2) Isolation of the protein - micro-organisms grown in huge fermenters and need to be isolated from the material on which they grow
3) Protein has to be purified to ensure it is uncontaminated
4) Microbial biomass can have a high proportion of nucleic acids which must be removed
Disadvantages of using micro-organisms in food (2)
5) Amino acid profile may be different from traditional animal protein - particularly deficient in methionine
6) Infection - conditions needed for micro-organisms to grow are also ideal for pathogenic organisms. Care must be taken to ensure the culture is not infected with the wrong organisms
7) Palatability - protein does not have the taste/ texture of traditonal protein sources
Fermenter
Commercial drug production can be controlled to ensure the best possible yield of the product
What must be controlled in the fermenter
1) Temperature - too hot the enzymes will denature and growth will be limited
2) Nutrients available - microorganisms require nutrients to grow and synthesise the product. Sources of carbon, nitrogen, minerals and vitamins are needed
3) Oxygen availability - most microorganisms respire aerobically
4) pH - enzyme activity and hence growth and synthesis are affected by extreme of pH
5) Concentration of product - if product is allowed to build up it may affect the synthesis of the product
How is a fermenter sterilised
Using superheated steam
What are the different parts of the fermenter
1) Pressure vent prevents any gas build up
2) Air inlet - sterile air provides oxygen in aerobic fermenters
3) Mixing blades
4) Water jacket inlet - allows circulation of water around the fermenter to regulate temperature
5) Outlet for draining the fermenter
6) Air outlet often in a ring - air bubbles out from outlets mix with culture
7) Electronic probes to measure pH/O2 / temperature
8) Water jacket outlet
9) Inlet for the addition of nutrients
10) Motor - rotates the blades so culture is mixed evenly
What are all inlets and outlets fitted with
Filters to prevent contamination
Primary metabolites
Products synthesised by the microorganisms during normal metabolism
-Continually released from the cells and can be extracted continuously from the fermenting broth
-Broth is topped up with nutrients as these are used by the microorganisms
- Some of the broth is removed - otherwise population becomes too dense
What kind of culture is primary metabolites
Continuous culture - keeps the microorganisms at a specific growth rate
Secondary metabolites
Products produced when the cells are placed under stress such as high population density or limited nutrient availability
-Produced mainly in stationary phase
-Culture is set up with a limited quantity of nutrients and allowed to ferment for a specific time
-Fermenter is then emptied and the product can be extracted from the culture
What kind of culture does secondary metabolites produce
Batch culture
Asepsis
Ensures sterile conditions are maintained
-Nutrient medium would also support the growth of unwanted microorganism which would reduce production
Asepsis - how would the growth of unwanted microorganisms reduce production
-Competition with the cultured microorganisms for nutrients and space
-Reduce the yield for useful products
-Spoil the product
-Produce toxic chemicals
-Destroy the cultured microorganisms and their products
Penicillin
Produced by mass fermentation of a fungus
-Secondary metabolite
-Produced by batch culture
Production of penicillin
1) Fermenter is run for 6-8 days
2) Culture is filtered to remove the cells
3) Antibiotic is precipitated as crystals by the addition of potassium compound
-Antibiotic may be modified by the action of other microorganisms or by chemical means
4) The antibiotic is mixed with inert substances and prepared for administration in tablet from / syrup for injection
Insulin
Obtained from a genetically modified bacterium by combing the human insulin gene with a plasmid to act as vector then inserting it into the E-coli
-Enabled the production of insulin by continuous culture
Bioremediation
The use of microorganisms to clean the soil and underground water on polluted sites
-Organisms convert toxic pollutants to less harmful substances
-Can break down crude oil/ solvents/ pesticides
-Stimulates the growth of microbes that use the contaminants as a food source
What are the right condition’s for microorganisms to grow (bioremediation)
-Available water
-Suitable pH
-Suitable temperature
What happens if conditions are not suitable (bioremediation)
Can be modified with the addition of certain substances
e.g. molasses/ pump O2 into aerobic bacteria
-Can be dug up and removed to be treated ex situ if conditions are not suitable in situ
Advantages of bioremediation
-Uses natural systems
-Less labour/equipment is required
-Treatment in situ
-Few waste products
-Less risk of exposure to clean up personnel
Disadvantages of bioremediation
Only suitable for certain products: heavy metals and lead cannot be treated
