Cloning and Biotechnology

Question 1

What is Biotechnology

Answer 1

Using an organism or parts of an organism in industrial processes

Question 2

What is a clone

Answer 2

Genetically identical organism or cell

Question 3

What are the methods of natural cloning by plants

Answer 3

Vegetative propagation - Plant body is separated then develops into a new plant

Runners - Horizontal stems that grow on the surface of the ground and can produce roots

Rhizomes - Horizontal stems that grow but underground

Suckers - New stems that grow out of the roots of a plant these parts are separated off to form a new plant

Bulbs - Overwintering mechanism, it consists of an underground stem from which can grow a series of fleshy leaf bases

Tubers - Another type of underground stem for example potato’s. One potato will grow into one of more plants

Question 4

What are the methods of natural cloning by animals

Answer 4

Formation of twins by embryo splitting. The zygote splits and it’s called monozygotic twins

Question 5

What are the advantages of natural clones

Answer 5

+ If conditions are favourable for growth of the parent they will also be good for the offspring
+ Cloning is rapid so population size can increase quickly
+ No need for a mate to carry out this reproduction

Question 6

What are the disadvantages of natural clones in plants

Answer 6

• The offspring may become overcrowded
• There will be no genetic diversity so there is susceptibility to disease
• Evolution is much less possible due to lack of variation

Question 7

What methods are there for artificial cloning of plants

Answer 7

Cuttings
Tissue culture
Micropropagation

Question 8

How would you take a cutting

Answer 8

Cut stem at a slant from a healthy plant
Dip in rooting hormone/powder
Place in watered compost
Place a bag on top to reduce transpiration

Question 9

What is the technique for micropropagation

Answer 9

Take some meristematic tissue known as an explant
Place in nutrient growth medium and sterilise with ethanol
Leads to a formation of a callus
Separate then place in shooting powder then rooting powder
Put in a greenhouse

Question 10

What are the advantages of artificially cloning plants

Answer 10

+ Rapid method compared to planting seeds
+ You can obtain desirable characteristic resulting in a high yield
+ Cn be carried out when sexual reproduction isn’t possible
+ Can be done any time of the year

Question 11

What are the disadvantages of artificially cloning plants

Answer 11

• Can be expensive to set up facilities
• No genetic variation unless introduced by mutation
• Genetically identical so susceptible to disease
• Tissue culture can fail due to contamination

Question 12

What are the three main types of animal cloning

Answer 12

Embryo splitting
SCNT
Therapeutic cloning

Question 13

What happens in embryo twinning

Answer 13

A zygote is produced in IVF
Allowed to divide by mitosis into small bags of cells
Separated and allowed to develop
Each mass is placed in a surrogate mother

The genotype and phenotype will depend on the sperm and egg used

Question 14

Describe the process of SCNT

Answer 14

Take a somatic cell from the animal you want to clone and remove the nucleus
Take an egg cell and remove the nucleus (enucleation)
Fuse the nucleus (from somatic cell) and egg cell and electro fuse it stimulating cell division
Place the young embryo in a surrogate mother

Question 15

What is therapeutic cloning

Answer 15

It’s non-reproductive and tissues and organs could be grown as replacement parts from the patients own cells. For example growing beta cells

Question 16

Why can clones be used for research

Answer 16

As they are genetically identical they will have the same effects as the cloned species. Drugs and medicines can be tested for its effects

Question 17

What the arguments for artificial cloning

Answer 17

+ Endangered species can be cloned to increase population size
+ Obtain desirable characteristics for high yield
+ Can be used for research

Question 18

What the arguments against artificial cloning

Answer 18

• Lack of genetic variation susceptible to disease
• Success rate is very low and even if successful clones have a shorter lifespan
• Ethical issues
• Cloning doesn’t increase genetic diversity

Question 19

How is yoghurt made

Answer 19

It’s milk that has undergone fermentation by Lactobacillus and Streptococcus. The bacteria converts lactose into lactic acid. This acidity denatures milk proteins causing it to coagulate. The bacteria partially digests the milk.

Question 20

How is cheese made

Answer 20

Lactobacillus produces lactic acid from lactose. The the milk is mixed with rennet (can now be GMO’d) which contains the enzyme rennin. Rennin coagulates the milk protein (casein). The casein gets broken down then is precipitated and separated from whey leaving its solid form of curd. The curd is pressed into moulds. During this stage different treatments can occur to determine the characteristics of cheese. For example in blue cheese inoculation with fungi gives additional flavour.

Question 21

How is bread made

Answer 21

Mixture of flour, water, salt and yeast called Saccharomyces cerevisiae. When this mixture is left to prove the yeast respires anaerobically producing CO2 making the dough rise. The alcohol evaporates. The toxic build up causes the yeast to die.

Question 22

How are alcoholic beverages made

Answer 22

It’s the product of anaerobic respiration in yeast. In wine making the crushed grapes provide sugars that yeast use to produce CO2 and alcohol. Ale or beer uses barely grains that are beginning to germinate. It converts stored starch to maltose which is respired by the test. Hops are used to give a bitter taste

Question 23

How is mycoprotein made what are the + and -

Answer 23

Fusarium venenatum is used to produce Quorn. It contains no animal fat or cholesterol but does have a high purine content so could lead to gout.

+ Faster process
+ High protein content
+ No animal welfare issues
+ Not much land required

• Don’t taste nice
• Protein has to be purified to ensure it’s uncontaminated
• Some people don’t want to eat food that’s been grown on waste

Question 24

What is a fermenter, what does it consist of and what factors need to be controlled

Answer 24

Used to grow microorganisms, conditions are controlled to ensure the best possible yield of product.
Has a pressure vent to prevent gas build up
Has a water jacket to take in heat and cool the vessel
An air inlet to provide sterile O2
Motor and paddles to give an even distribution
A probe to monitor conditions

Temperature - Don’t want enzymes to be denatured
Nutrients - Needed for nourishment
Oxygen - Aerobic respiration
pH - Needed for enzymes

Question 25

What is the typical growth phase of an microorganism

Answer 25

Lag phase - Switching genes on and acclimatising to conditions
Log phase - Population size doubles there are plentiful resources og*2^n
Stationary phase - Nutrient levels decrease , waste metabolites build up death=reproduction
Death phase - All nutrient used up, toxic conditions

Question 26

What is a primary metabolite

Answer 26

Primary metabolite are products that are only synthesised during normal metabolism when microorganisms are actively growing. As these products are continuously released they need to be extracted continuously. The procedure is maintained by topping up with nutrients. The culture can be maintained in the log phase to gain high yield. An example would be mycoprotein.

Question 27

What is a secondary metabolite

Answer 27

Products produced when cells are placed under stress such as limiting nutrients in the stationary phase. For this reason nutrients are usually only placed in at the start and the product is collected when the fermentation is emptied. This is known as batch culture and a common example is penicillin produced by penicillium when under stress.

Question 28

Compare batch and continuous culture

Answer 28

Batch culture is easy to set up but growth rates are slower due to build up of toxins
Only the batch is lost if the fermentation becomes contaminated
You can continually remove product in continuous and quality is more consistent
In batch culture the down streaming process is much longer in between fermentations

Question 29

How can micro organisms be genetically modified and what are the benefits

Answer 29

You insert the gene used to produce insulin into a bacteria such as E.coli
Previously insulin was extracted from pigs but this method ensures there are no allergies and ethical issues as the insulin produced is human insulin.
Vast quantities can be produced by continuous culture

Question 30

What is bioremediation

Answer 30

The use of microorganisms to clean soil and underground water on polluted sites. Toxic pollutants are converted into less harmful substances.
An example is GM Pseudomonas bacteria that can break down crude oil from oil spills.
In some cases UV light can be used to active the genes needed to break down substances

+ Less labour
+ Fewer waste products
+ Treatment is in situ

• Only suitable for certain products

Question 31

How would you go about growing microorganisms on agar

Answer 31

Sterilisation - Heated to 121 degrees in an autoclave to kill any living organisms
Inoculation - (how microorganisms are introduced to medium) methods include streaking, seeding (sterile pipette transfers medium onto agar surface) and spreading (sterile glass spreader evenly distributes inoculated drop)
Ethanol is used to sterilise equipment
Lid is opened slightly to stop unwanted microorganisms settling and a Bunsen burner is nearby to waft and bacteria away

Question 32

How should you incubate your agar plate

Answer 32

Place upside down to stop condensation and prevent the agar from drying out too quickly
The temperature is set at 25 degrees to stop unwanted microorganisms growing on surface

Question 33

How could you determine the number of viable colonies on an agar

Answer 33

Serial dilutions can be performed in order to reduce the concentration of solution or suspension. Each following dilution is always a 1/10 of the previous one. You would do this about 5 times then place each culture on agar plates looking for results that show countable colonies.
Then you get the dilution common factor (DCF) and scale up to the original sample

E.g. 18 colonies in 100 microliters then in that 1ml sample there is 180 per ml and if that was 10^-4 you would times by 10^4 to obtain colonies in original sample

Question 34

What are immobilised enzymes

Answer 34

They are held in place and aren’t free to diffuse through the solution

Question 35

What are the advantages of immobilised enzymes

Answer 35

Fewer unwanted products from reaction
Shorter downstream process as product is already separated
Protected from extreme temperatures and pH
These enzymes can be reused

Question 36

What are the 4 main ways of using immobilised enzymes

Answer 36

Adsorption - Held by hydrophilic/phobic interactions and ionic bonds, adsorbed onto inert material
Covalent bonds - Held by covalent bonds
Membrane separation - On one side of a partially permeable membrane and substrate on the other side
Entrapment - Enzyme trapped in porous bead for example algal balls

Question 37

What are the disadvantages of immobilised enzymes

Answer 37

These are expensive
Chance of leaking
Active sites can get blocked

Question 38

What are some industrial uses of immobilised enzymes

Answer 38

Glucose isomerase - Glucose to fructose (used as a sweetener)
Lactase - Lactose to glucose and galactose
Penicillin Acylase - Forms semi synthetic pennillions, some resistant bacteria aren’t resistant
to these
Aminoacylase - Hydrolase to produce pure samples of L amino acids
Glucoamylase - Converts dextrins to glucose, used to sweeten and thicken foods