6.2.1 Cloning and biotechnology

Question 1

What is a Clone?

Question 2

What is vegetative propagation?

Answer 2

A form of asexual reproduction where new, genetically identical individuals develop from non-reproductive tissue of parent plant, such as its roots, leaves & stems.

Question 3

What is asexual reproduction?

Answer 3

The production of genetically identical offspring (clones) from a single parent, without the fusion of gametes.

Question 4

What are the 5 methods of natural vegetative propagation?

Answer 4

• Rhizomes
• Stolons or runners
• Suckers
• Tubers
• Bulbs

Question 5

What are rhizomes in vegetative propagation? Give an example.

Answer 5

Rhizomes are stem structures that grow horizontally underground away from parent plant.
They have ‘nodes’ which new shoots and roots grow from.
Example = Bamboo or Marram grass

Question 6

What are stolons (runners)? Give an example.

Answer 6

Similar to rhizomes, horizontal stems that grow along soil surface away from the parent plant, with nodes or stem tips that can root to form a new plant.
Example = Strawberries

Question 7

What are suckers in plants? Give an example.

Answer 7

Shoots that grow from sucker buds (undeveloped shoots) present on the shallow roots of parent plant.
Example = Elm trees

Question 8

How do tubers function in vegetative propagation? Give an example.

Answer 8

Tubers form when a stem tip swells with food, with buds on the surface that develop into new shoots.
Example = Potatoes

Question 9

What are bulbs in vegetative propagation? Give an example.

Answer 9

Bulbs form when a leaf base becomes swollen with stored food, the inner bud can form new shoots.
Example = Onions or daffodils

Question 10

Outline an experiment to artificially propagate plants from cuttings from a stem.

Answer 10

1. Cut a 5-10 cm piece from the end of a parent plant’s stem using a sharp, sterile tool.
2. Remove the lower leaves, leaving only one leaf at the top.
3. Dip the cut end in rooting powder, which contains plant hormones that encourage root growth.
4. Plant the cutting in a suitable growth medium, such as compost.
5. Place it in warm, moist conditions to promote root development.
6. Once rooted, transplant the new clone.

Question 11

How would you artificially propagate root cuttings or leaf cuttings?

Answer 11

• Root cuttings - Take a section of root and make an angled cut on one end before treating it as you would a stem cutting.
• Leaf cuttings - Remove an entire leaf, score the veins, and place it in a growing medium with the scored veins facing down.

Question 12

List some advantages of vegetative propagation.

Answer 12

• Fast and cost-effective.
• Ensures a high yield
• It maintains quality of the crop because the new plants have same genetic traits
• Allows survival in adverse conditions and regeneration each season

Question 13

List some disadvantages of vegetative propagation.

Answer 13

• Lack of genetic variation in offspring
• More susceptible to diseases, pests and climate change

Question 14

What is micropropagation?

Answer 14

A technique used for producing many identical plant clones from a single parent plant through tissue culture.
It is a type of asexual reproduction on a very large scale.

Question 15

What is tissue culture?

Answer 15

Involves growing plant tissues in a sterile medium enriched with hormones (auxins and cytokinins) that stimulate cell division and growth.

Question 16

What are the five key steps of tissue culture?

Answer 16

1. Explant collection
2. Sterilisation
3. Culture
4. Development
5. Transfer

Question 17

What is an explant and why is it used in micropropagation?

Answer 17

An explant is a small tissue sample taken from a parent plant, usually from root or stem tips where meristem cells are present.
These cells are totipotent, meaning they can differentiate into any plant cell type.

Question 18

Why is sterilisation important in micropropagation?

Answer 18

To remove and inhibit the growth of unwanted microoganisms, such as bacteria and fungi
Reduces infection risk and ensure healthy plant development.

Question 19

What does the culture medium provide in micropropagation?

Answer 19

Nutrient-rich medium.
Supplying mineral ions, sugars, vitamins and growth hormones (auxin & cytokinin) to encourage shoot and root formation.

Question 20

What is a callus in micropropagation?

Answer 20

A callus is an **undifferentiated **mass of plant cells formed when explant cells divide.
It is later transferred to a new medium with specific conditions to differentiate into shoots and roots and soon plantlets.

Question 21

What happens in the final stage of micropropagation?

Answer 21

Fully developed plantlets are moved to soil or a growth medium to grow into mature plants, genetically identical to the parent.

Question 22

Name three applications of micropropagation.

Answer 22

• Rapid propagation of endangered or slow-growing plants
• Production of disease-free or GM clones
• Production of seedless plants or those that are difficult to cultivate from seeds

Question 23

Give some advantages of micropropagation.

Answer 23

• Desirable genetic characteristics are always passed on to clones
• Plants can be produced in any season as environment is controlled
• Less space required compared to conventional farming methods
• Rapidly produces of a large number of mature plants

Question 24

Give some disadvantages of micropropagation.

Answer 24

• Lack of genetic diversity - monoculture meand higher vulnerability to disease
• Very high production costs due to high energy costs and training required
• Contamination by microogranisms can be disastrous and result in complete loss of culture

Question 25

What is **natural** cloning in animals?

Answer 25

Occurs when animals produce genetically identical offspring using asexual reproduction.

Question 26

How does natural cloning occur in vertebrates👶?

Answer 26

An early embryo splits into two genetically identical embryos. Each embryo grows independently.

Question 27

What is artificial **embryo twinning**?

Answer 27

A method where an early embryo is **manually split** into individual cells before differentiation to produce genetically identical embryos.

Question 28

Outline the steps in artificial embryo twinning in cows.

Answer 28

1. An **egg cell** is extracted from a female cow and fertilised in a Petri dish 2. The fertilised egg is left to divide, forming an **embryo** *in vitro*. 3. The individual cells from the embryo are **seperated** and each is put into a seperate Petri dish. Each cell divides and develops normally, so an embryo forms in each dish 4. Embryos are then impanted into female cows (surrogates). 5. Embryo develops in womb of cows, and eventually birthed - all offspring will be **gentically identical**.

Question 29

What is **somatic cell nuclear transfer** (SCNT)?

Answer 29

A technique where a **nucleus** from a **somatic cell** is inserted into an **enucleated** ovum, creating a clone of the donor.

Question 30

Outline the steps in somatic cell nuclear transfer in sheep.

Answer 30

1. A **somatic cell** is taken from sheep A. The nucleus is extracted and kept. 2. An **oocyte** (immature egg cell) is taken from sheep B. Its nucleus is removed to form an **enucleated oocyte**. 3. The nucleus of from sheep A is inserted into the enucleated oocyte - the oocyte from sheep B now contains the genetic info from sheep A. 4. The nucelus and enucleated oocyte are **fused** together (**electrofusion**) and stimulated to divide - producing an embryo. 5. Embrto is implanted into suurogate mother and eventually a lamb is born - a clone of sheep A.

Question 31

Name some applications of animal cloning.

Answer 31

* **Medical research** - Cloning produces genetically identical animals for drug testing and disease modelling. * **Conservation** of endangered species from a limited gene pool. * **Agriculture** - Cloning can replicate animals with desirable characteristics for selective breeding to improve product quality. * **Pharming** - Genetically engineered animal clones can be used to produce therapeutic proteins. * **Embryonic stem cells** - Replace damaged tissues.

Question 32

✅Give some arguments **in favour** of animal cloning.

Answer 32

* Desirable characteristics are always passed on (e.g. high milk yield) * Infertile animals can be reproduced * Increase pop. of endangered species to preserve biodiversity. * Facilitates medical advancements and new treatments for diseases that could **alleviate suffering**.

Question 33

❌Give some arguments **against** animal cloning.

Answer 33

* Cloning is difficult, time-consuming & expensive * No genetic variation, undesribale characteristics (e.g. weak immune system) are always passed on, so all clones are susceptible to the same diseases. * Potential for shorter lifespans in clones. * Ethical concerns regarding the destruction of embryos. * Low success rates and health issues in clones.

Question 34

What is **biotechnology**?

Answer 34

The industrial use of living organisms or their components (enzymes) to produce food, drugs and other products.

Question 35

What type of living organism are mostly used in biotechnology?

Answer 35

Microorganisms Such as bacteria, fungi, moulds & algae.

Question 36

What are some reasons for the use of microogranisms in biotechnology?

Answer 36

* Their ideal growth conditions can be **easily** created. They will grow successfully as long as they have the right nutrients, temperature, pH, moisture levels and gases. * Have a short-life cycle so **grow rapidly** under right conditions. * Grown **any time** of the year.

Question 37

List some applications of microbes in biotechnology.

Answer 37

* Brewing * Baking * Yoghurt * Medicines

Question 38

Give some advantages of using microorganisms in biotechnology.

Answer 38

* Microorganisms can be grown quickly, easily & cheaply * Less land is required in comparison to growing crops or rearing livestock * Many microorganisms can grow and reproduce using waste materials as their nutrient source. * No animal welfare issues * High protein and low fat content make them an efficient food source

Question 39

Give some disadvantages of using microorganisms in biotechnology.

Answer 39

* Sterile conditions are necessary, increase operational costs * Risk of contamination by unwanted microorganisms as condition are ideal for other * Taste/texture may differ from traditional foods * Seperation of microorganisms from nutrient broth is required for food production

Question 40

What are primary and secondary metabolites in microbes?

Answer 40

* **Primary metabolites** are substances produced in processes that are essential for normal microbial function. (e.g. ethanol). * **Secondary metabolites** are produced later, in non-essential processes (e.g. antibiotics).

Question 41

🧫 What is a bioreactor/fermentation vessel?

Answer 41

A large fermentation tank that creates **optimal conditions** for microbial growth to produce biomass or desired products.

Question 42

How are conditions optimised in a fermentation vessel?

Answer 42

* **pH** - Constantly monitored by a **pH probe** and kept at the optimal level, allows enzymes to work efficiently, maximises rate. * **Temperature** - Kept constant by a **water jacket** surrounding entire vessel, allows enzymes to work efficiently, maximises rate. * **Nutrient access** - **Paddles** constantly circulate fresh nutreint medium around vessel, ensures microbes always have access to required nutrients. * **Vol of Oxygen** - Sterile air is pumped into vessel when needed, for respiration. * **Sterile conditions** - **Superheated steam** sterilises vessel after each use, kills unwanted organisms that may compete with culture.

Question 43

What’s the difference between **batch** and **continuous** fermentation?

Answer 43

* Batch fermentation - Microorganisms grown in individual batches, fixed volume, nutrients run out, culture is replaced after each batch. * Continuous fermentation – Microorganisms continually grown in vessel without stopping, fresh nutrients added continuously, product and waste removed constantly.

Question 44

What are the four phases of microbial growth in batch fermentation? (refer to the graph)

Answer 44

1. **Lag phase** - Cells have slow initial growth as they adapt to their environment and produce essential enzymes. 2. **Log** (**exponential**) **phase** - Rapid doubling of cell numbers occurs under ideal conditions, and growth rate is at its maximum. 3. **Stationary phase** - Growth rate plateaus as nutrients diminish and waste accumulates, and cell growth is **equal** to cell death. 4. **Death phase** - Cell death rate exceeds cell growth rate due to resource limitation and build up of **toxins**.

Question 45

How can you work out how many individuals will be present in a population after a number of division?

Answer 45

**N = N₀ × 2ⁿ** N = final population size N₀ = initial population size n = number of divisions

Question 46

Outline the steps to culture microbes on agar plates.

Answer 46

1. **Sterilise** equipment (e.g. flame the inoculating loop). 2. Using a sterile pipette, add a set vol. of a given sample of bacteria in broth to an agar plate. **Discard** pipette safely after use. 3. Spread the broth across the entire surface of the agar using sterile plastic spreader and discard safely. 4. **Repeat** step 1-3, so that there are 6 plates total 5. Seal plate **lightly** with tape (to allow air exchange). 6. Label plate with microbe type, date, and conditions. 7. Incubate **upside down** at appropriate temp. - stops condensation forming on lid and dropping onto agar. 8. Repeat as control using a sterile loop with no microbes - negative controls. 9. Assess growth by observing bacterial colonies.

Question 47

Why is a control plate used?

Answer 47

It ensures that the agar itself is **sterile** (no contamination) Any microbial growth seen on experimental plates is due to the **added bacteria**, not from airborne microbes or contaminated equipment

Question 48

Why are the agar plates incubated upside down?

Answer 48

Prevents condensation forming on the lid and dropping onto the agar. This could cause contamination.

Question 49

What are some control variables in an agar plate microbial culture?

Answer 49

Temperature Incubation time Type and volume of nutrient agar Volume and strain of bacteria used Sterilisation method and inoculation technique pH of the medium Oxygen availability

Question 50

What **aseptic techniques** are used when culturing microorganisms?

Answer 50

- Regulary disinfect work surfaces to minimise contamination - Work near a bunson burner creating a convection current that carries microbes away - Sterilise instrument used to transfer cultures before and after, e.g pass wire innoculating loop & spreader by passing it through flame. - Flame the neck of the bottles of bacteria or broth -.Minimise time that agar plate is open and put lid on as soon as possible - Wear a lab coat, gloves, and tie long hair back.

Question 51

Name four factors that affect microbial growth and how they can be investigated.

Answer 51

- **Temperature** - Incubate duplicate plats at different temperatures - **pH** - Add buffers to the agar to maintain different pH levels - **Nutrient availability** - Prepare agar with varying conc. - **Antimicrobial resistance** - Add different antimicrobial compounds to agar plate.

Question 52

What are immobalised enzymes?

Answer 52

Enzymes that are attached to an insoluble material so they cannot be mixed with products.

Question 53

Why is enzyme immobilisation important in industry?

Answer 53

Isolated enzymes can become mixed in with products of reaction which have to then be **separated** from mixture. Enzymes are expensive, immobilisation allows their reuse, reducing costs and improving stability.

Question 54

Name three main methods of enzyme immobilisation.

Answer 54

1. **Encapsulated** in jelly-like alginate beads, which act as a semi-permeable membrane. 2. **Trapped** in a silica gel matrix 3. **Covalently bonded** to cellulose or collagen fibres.

Question 55

What are the advantages of using immobilised enzymes?

Answer 55

* Reusable enzymes reduce costs * Enzyme-free products improve purity, less money & time spent on separation. * More stable under temperature and pH changes, they are less likely to denature.

Question 56

What are the disadvantages of immobilised enzymes?

Answer 56

* Higher initial costs for materials and extra equipment * Possible reduced enzyme activity * More complex reactor systems prone to technical issues

Question 57

Name some industrial processes that use immobilised enzymes.

Answer 57

- Conversion of lactose to glucose & galactose for lactose-free milk - Immobalised lactase. - Production of semi-synthetic penicillins - Immobalised penicillin acylase. - Conversion of glucose to fructose (much sweeter) - Immobalised glucose isomerase.

Question 58

How is immobilised **lactase** used to produce **lactose-free milk**?

Answer 58

1. Lactase is immobilised on alginate beads packed into a column. 2. Milk flows through, lactose is broken down into **glucose** and **galactose**. 3. Immobilised lactase stays in column to process more milk continuously.