6.4 Cloning And Biotechnology Flashcards
What are clones
Genetically identical organisms or cells
How can clones be produced
asexually (mitosis, budding, binary fission) or sexually
Advantages of natural cloning
if the condition is good for the parents it will be good for the offspring = high yield of the product, cloning is fast, reproduction can happen even in the absence of parents = good if it’s a rare species
Disadvantages of natural cloning
through natural cloning areas can become overcrowded by offspring, no genetic diversity unless there’s mutations = little variation, if something happens that wipes one of them out it’ll wipe all of them out
What does plant cloning happen using
Their vegetative parts = aka vegetative propagation = natural cloning in plants involves this
Examples of how vegetative parts of plant are creating clones:
Runners/stolon, suckers
Bulbs, corms
Tubers
how natural cloning occurs in plants bc it gives rise to new plants that are genetically identical.
Runners/stolon
when the stems grow on the surface of the ground
Rhizome
If the stems grow underground
Sucker
extension of the roots underground
Tubers
underground stems e.g. potatoes = example of plant natural cloning
Natural cloning in animals overview
When zygote splits into 2 daughter cells + produces twins
Natural cloning in insects
not common but water fleas and green flies can reproduce asexually.
Plant cuttings
- to get a plant cutting you have to cut the stem at two nodes (internode contains meristem tissues)
- The stem should then be placed in moist soil and new roots will form
- Some plants will naturally form roots, some may need help so you can add rooting powder (contains rooting hormones)
Where else can plant cuttings be taken from
Root, leaf
Larger scale version of plant cuttings
Tissue culture (vegatative propagation
Vegetative propagation
concept of growing cells, tissues or organs from a small sample of cells/tissues
To make tissue culture large scale
Micro propagation
Example of piece of plant tissue is used for Micropropagation
a leaf (best piece to use is meristem because it is free from viruses)
Callus
small lump of undifferentiated cells
Outline Micropropagation
-take a small piece of plant tissue
- Cut this into smaller pieces + call these pieces the explants
- Take the explants and sterilise them using bleach or alcohol
- Place the explant on a sterile growth medium e.g. agar containing suitable nutrients including glucose, amino acids, auxins and cytokinins = stimulates mitosis in the plant so you’ll see some growth = initial growth is known as a callus
- Take the callus and move it to a new agar which has a nutrient content to promote shoot growth = needs high concentration of auxins compared to cytokinins
- Once the shoot grows move it to a new agar which has nutrient content to promote root growth = needs low concentration of auxins compared to cytokinins = gives rise to tiny plants called plantlets.
- Move plantlets to a greenhouse to be grown in soil/compost
Advantages of artificial cloning in plants
- Fast
- Can be done when sexual reproduction is not possible
- Plant would have genetically identical features to parents = if these are desirable characteristics e.g. pest resistant or disease resistant, this can be beneficial
- Plants have the same phenotype = easier to grow + harvest them together
- When using meristems = new plants are free from viruses
Disadvantages of artificial cloning in plants
- labour intensive
- Expensive to set up the ideal conditions = specialised equipment
- Low genetic diversity = all clones are susceptible to same diseases
- No genetic diversity unless there’s a mutation
Use of reproductive cloning
for selective breeding, to increase the yield of a good characteristic
2 main techniques of reproductive cloning
Embryo twinning and somatic cell nuclear transfer
Embryo twinning
- fertilise the egg with a sperm in vitro of two high value animals of the same species
- Let zygote divide by mitosis to form small balls of cells
- Impregnate surrogate mothers with small balls of cells
Somatic cell nuclear transfer
- Take body cells from the sheep you want to clone and take an egg donor
- Remove the nucleus from both the egg and the body cell and place the desired nucleus into the empty egg cell
- Fuse them together using electrofusion
- Put the new egg cell into the sheep so that it starts replicating and collect the early embryo
- Implant the early embryo into a surrogate mother
- This clones the exact body cell’s phenotype
non reproductive cloning in animals
- process of cloning cells/tissues but not the whole animal
What can you do with non reproductive cloning in animals
- Can grow skin in a lab = can be used to repair damage
- Can be used for medical damage e.g. spinal chord issues, restoring the function of the pancreas
- Can grow organs in the lab
Benefit of non reproductive cloning in animals
less chance of rejection bc it’s the patient’s own body cells
Advantages of non reproductive cloning in animals
- can produce high yield of ideal characteristics
- Genetically identical embryos can be used for scientific research to check the effect of environment on a gene
- Drugs can be tested on cloned cells and tissues to avoid testing on humans/animals
- Therapeutic cloning reduces the chances of rejection
- Individuals from endangered species can be cloned to increase their numbers
Disadvantages of non reproductive cloning in animals
- lack of genetic variation = all susceptible to the same disease
- Success rate is poor and the method is very expensive
- Ethical issues regarding how long the embryo survives
- Doesn’t increase genetic diversity
Biotechnology
- use of microbes/ any living thing for commercial purposes
Most common uses of biotechnology
for food and job production
Why are microbes used in biotechnology
fast growth rate, no growth requirements
Examples of microbes in biotechnology (food)
Organism used:
- yeast to make ethanol for beer
- yeast as source of carbon dioxide for baking
- lactobacillus to make lactic acid for yoghurt + cheese
- mycoprotein as a source of protein for vegetarians
What is mycoprotein made from
Fungus
Microbes in Biotech examples (pharmaceutical drugs)
penicillium fungus for penicillin, other antibiotics and insulin
Microbes in Biotech examples (enzymes)
protease and lipase in washing powders uses enzymes taken from bacteria
lactase from bacteria to make lactose free milk
Bioremediation
bacteria and fungi can be used to cleanse water
ADVANTAGES OF USING MICROBES IN BIOTECH:
- cheap + easy to grow
- Fast reproduction rate
- Production takes place at lower temperatures and pressures than normally required
- Production isn’t climate dependent = can happen anywhere in the world
- Can use byproducts from other reaction as nutrients for bacteria and fungus in these reactions
- Easier to genetically modify microbes
- Few ethical considerations
- Easier to purify + isolate products for microbes than in the conventional method
Other forms of biotechnology
- gene technology
- Genetic modification + gene therapy
- Selective breeding
Examples of using biotech to make food
Brewing, baking, cheese, yoghurt
What is brewing and what does it use and why
alcohol production + uses yeast = promotes anaerobic respiration in yeast. Grapes used bc grapes have yeast on their surface + also contains glucose and fructose
Explain brewing using grapes
When the grapes are crushed, the Yeast on surface uses sugars from the grapes to anaerobically respire to produce carbon dioxide and alcohol
Explain brewing for beer
use barley grains just at the peak of germination = as the grains start to germinate they convert starch into maltose = used by yeast to respire anaerobically to produce co2 and alcohol.
Baking
yeast used = proof dough to allow the yeast to respire anaerobically = produces the co2 that allows the dough to rise.
Cheese
milk is treated w bacteria that uses lactose to make lactic acid = acidifies the milk. Milk is then mixed with rennet which contains the enzyme rennin. Rennin coagulates milk under the presence of calcium ions= forms curd which is separated from the liquid components of milk. The bacteria keeps forming more lactic acid. Take the curd and press it into moulds = produces cheese. Microbes can also be used to flavour cheese.
Whic bacteria ferment milk for yoghurt production
lactobacillus and streptococcus thermophilus
Yoghurt production
Allow bacteria to ferment milk
Lactose converted to lactic acid
Denatures the milk proteins which allows the milk to coagulate. Bacteria partially digest the milk = easier to digest yoghurt. Fermentation process is what gives rise to the taste of yoghurt. Probiotics can be added to yoghurt from other microbes to make it more nutritionally beneficial.
What else can biotechnology be used to make
Penicillin and insulin
Biotech for penicillin
secondary metabolite of the fungus penicillium
Biotech for insulin
genetically modify E. coli bacteria to produce human insulin
Bioremediation
- when microbes are used to clean soils and underground water
What does bioremediation do
Convert toxic pollutants to less harmful substances
Example of bioremediation
pseudomonas = treats oil spills = need to create the right conditions for the microbes to use the pollutants as a food source.
How do you create the right conditions for the microbes to use the pollutants as a food source
by changing the temperature, the pH, adding additional nutrients.
Advantages of bioremediation
- use a natural system
- less equipments used
- fewer waste products
- less risk of exposure to the workers
- treatment happens in situ
Is bio remediation done in situ or ex situ
can be done in situ, ex situ depending on if the conditions are favourable in situ
Quorn
Aka single celled proteins
fungus produces single celled proteins e.g. mycoprotein acts as meat substitute.
Advantages of using fungal protein (quorn)
- faster to use plant (fungus) proteins
- Production can either increase or decrease to demand
- No animal killing
- No animal fat or cholesterol
- Can genetically modify proteins to increase it’s content of amino acids
- Not much land is required
Disadvantages of using fungal protein (quorn)
- some people don’t want to eat fungal proteins
- Isolation process needed to separate protein from fungus = has to be purified = extra step
- Can cause infections
- Although it’s a good source of protein, it’s amino acid profile will be different to that of protein
What is used for the mass production of quorn
Fermenters
Why is a fermenter used for mass production of quorn
- Can control its conditions e.g. temperature, nutrient availability, oxygen availability, concentration of product
What’s the most important step for mass production of quorn
to sterilise the fermenter using a rlly high temperature initially.
What else is used in the mass production of quorn and why
- pH probe = monitor pH to maintain optimal conditions for the growth of whatever microorganisms you want
- Water jacket = temperature control
- Air inlets = to provide the right amount of oxygen
- Inlets for nutrients too
2 types of culture
Batch and continuous
Continuous culture
continuously promote the growth of your product = keep adding nutrients + remove some product + keep adding nutrients. Remove product bc product build up could be toxic.
Batch culture
culture set up with limited quantity of nutrients + allow it to ferment for a while and then take out the product after the set time
Why is using batch and continuous cultures important
bc microbes produce 2 types of products, primary metabolites and secondary metabolites
Primary metabolites
products that are released as part of the normal growth of a microbe = nothing special
Secondary metabolites
released in cases of threat/selection pressure to compete w other microbes e.g. antimicrobials and antibiotics
Importance of asepsis using fermenters
- provide a nutrient medium = could also promote the growth of unwanted microorganisms so then you’ll have the ones you want to grow and their competitors competing for nutrients and space = will have a low yield of ur product and a contaminated product + competitors may also start producing toxic products. THIS IS WHY STERILE CONDITIONS ARE IMPORTANT.
Why do you use a microorganism culture for viruses
- Can’t grow viruses in a lab because they need the machineries of a host cell = take a sample and put it through PCR
How are bacteria and fungus taken for microorganism cultures
Culture a swab
Two types of growth medium for microorganism cultures
can grow bacteria and fungus or an agar plate or nutrient broth (liquid medium)
Purpose of microorganism cultures having two growth mediums
they have nutrients that promote growth e.g. salts, sugars, some can have blood
Why are aseptic techniques
To prevent contamination
Aseptic techniques for microorganism cultures
- Wash your hands.
- Disinfect the working area.
- Have a Bunsen burner operating nearby to heat the air. This causes the air to rise and prevents air-borne microorganisms settling, It also creates an area around it of sterile air in which the microbiologist can work.
- As you open a vessel, pass the neck of the bottle over the flame to prevent bacteria in the air entering the bottie. The bottle should also be flamed as it is closed.
- Do not lift the lid of the Petri dish off completely - just open it enough to allow introduction of the desired microorganism.
- Any glassware or metal equipment should also be passed through the flame before and after contact with the desired microorganisms
How to sterilise microorganism cultures
Put the Petri dish and the inoculating loop through an autoclave at 121 degrees celsius to sterilise everything.
How to create agar
- open bottle over Bunsen burner flame
- When pouring agar gel and only open lid slightly, enough to pour the liquid into the plate
Inoculation
introduction of microorganisms to the sterile medium (taking bacteria from a liquid broth and spreading it over agar)
Different ways to inoculate?
Streaking, seeding, spreading
Streaking
bc broth has lots of bacteria inside, when performing streaking the aim is to isolate one bacterial colony to see its morphology
Difference between streaking and other techniques
Streaking isolates colony, other techniques allow you to grow a bacteria
Steps of streaking
Why is the agar lid slightly open
To prevent contamination
Seeding
a sterile pipette is dipped in the liquid broth and this is dropped over the surface of the agar
Spreading
using a sterile glass spreader to spread the bacteria from the liquid broth over the surface of the agar
What else can be used for inoculation instead of seeding or spreading
Can also use a moist cotton swab with distilled water and placing this over the agar and closing the agar
Incubation
keeping the taped up agar plate at a specific temperature for a set amount of time.
What should you do when closing the lid for incubation
Don’t tape it fully
some air movement to allow for the growth of some organisms. And make sure the Petri dish is labelled.
What should you do when storing the Petri dish for incubation
store it in a warm environment to allow for bacterial growth but store it upside down = prevents condensation from falling back onto the microorganisms
What do bacteria and fungus look like after an incubator is used
If it’s spongey + green = likely to be a fungus
Shiny circles = bacteria
Using a liquid medium
allows bacteria to grow but you won’t be able to identify characteristics of the bacteria but can measure population growth by seeing how much light passes through the broth
Serial dilution using streak plating
If there’s x amount in the smallest dilution you can work your way backwards to calculate how many were in the original broth culture.
= use serial dilution and plating.
Serial dilution using streak plating diagram
Units for counting bacteria
cfu (colony forming units)
Population growth curve in a closed culture
Lag phase
time taken for bacteria to acclimatise to the environment. E.g. cell growth, switching on genes, protein synthesis.
Exponential phase
when the bacteria are used to environment and start reproducing and there’s plenty of nutrients and space = no setbacks
Stationary phase
when the organisms have started to use up the nutrients and there’s more waste in the system now and the reproduction rate and death rate are equal
Death/decline phase
when organisms run out of nutrients and there’s a build up of waste so some organisms are killed.
When are primary metabolites made in terms of the population growth curve
made part of the natural growth and are normally made during the log phase
When are secondary metabolites made in terms of the population growth curve
only produced when the bacteria sense danger (when it wants to start competing with someone) e.g. antibiotics = produced late stationary phase/early decline phase.
Are antibiotics primary or secondary metabolites?
Secondary
How do you collect a secondary metabolite
- create a closed system w limited resources = induces stationary/decline phase environment
Immobilised enzymes
- new invention where enzymes are held in place and aren’t free in solution = can immobilise them to surfaces where they’re stuck to something.
Advantages of immobilising enzymes
- if you wanted to make a solution and needed an enzyme just pass a substrate through a mixture and the product will gather at the bottom.
- Enzymes aren’t used up in the reaction but they remain in the beaker = means at the end they’d need a purification step
- Benefits of this is that enzymes don’t mix with the products, so the purification step would be cheaper
- Easier to reuse the enzymes
- Only putting enzymes in rather than cells that make the enzymes = better bc all the requirements for the cell e.g. nutrients and reproduction stages would no longer be needed
- By immobilising the enzymes you are protecting the enzymes from extreme conditions = high temperature and pH can be used.
Methods for enzymes to bind to surface
Adsorption, covalent bonding, entrapment, entrapment w membrane separation
Adsorption
- Bind enzymes to a surface using hydrophobic interactions and ionic links.
- Aim is to stick the back of the enzymes to these surfaces so that the active site is exposed but because using ionic links, the shape of the active site might get slightly distorted.
What can enzymes bind to in adsorption
Clay, glass beads, resins
How can enzymes detach in adsorption
can detach from the surface because the hydrophobic interactions and ionic links are weak.
Covalent bonding
- enzymes use covalent bonds to bind to the clay particle and cross linking agents can be used to add more enzymes
Benefit and negative of covalent bonding
covalent bonds are strong so they are unlikely to come out. But because the bonds are strong they are more likely to distort the active site shape = reducing activity
Entrapment
- trap the enzyme in a matrix = enzymes aren’t directly exposed to the substrate = substrate and products have to go through the matrix and then come out of the matrix.
Why is entrapment safer for enzymes
because there’s no bonding involved which can alter the active site = only enclosed in matrix but the products and substrate would have to diffuse out of the matrix so they’d have to be smaller molecules = not suitable for all reactions.
Example of entrapment medium
Cellulose mesh
Entrapment a membrane separation
- membrane separating the enzymes and the product and substrate = the product and substrate will only pass through if they’re permeable to the membrane. = alternative version of entrapment
Industrial uses of immobilised enzymes
Glucose isomerase, penicillin acylase, lactase, aminoacyclase, glucoamylase
Glucose isomerase
- converts glucose to fructose, which is now used to make most of the sugary food because it a lot sweeter so you need less sugar
Penicillin acylase
- Form semi synthetic penicillin = to get ahead of antibiotic resistance
Lactase
- breaks down lactose to glucose and galactose
- Produce lactose free milk
Aminoacyclase
- Produces pure samples of L amino acids = can be used in the pharmaceutical industry to create compounds
Glucoamylase
- converts dextrins to glucose
What are viruses
Bits of genetic code
Do viruses have dna or rna
Some have dna, some have rna
Examples of viruses w rna
HIV, influenza and rabies
Examples of viruses w dna
Warts and chickenpox
Which virus is most likely to mutate + why
RNA viruses = more likely to mutate because there’s only one strand = risked
What do viruses rely on
Reverse transcriptase
What’s a virus’s aim
To get it’s genetic material inside a cell
Disadvantages of using microorganisms
Some people may not want to eat fungal protein or food grown on waste
• Need to isolate the protein from the medium it is grown in
• Protein has to be purified
• Culture can become infected easily due to optimum growing conditions
• Different taste/texture to traditional protein sources
Why can no population grow indefinitely
Limiting factors e.g.
Food
• Shelter
• Light
• Water
• Oxygen
• Accumulation of toxic waste
• Disease
• Predators
