Module 6 cloning and biotechnology

what is meant by cloning?

The process of using an organism to produce genetically identical cells / organisms.

How can cloning occur

Naturally ( Asexual reproduction)

Artificially

How do plants produce natural clones?

Vegetative propagation

what is Vegetative propagation?

Production of plant clones from non-reproductive tissues

what type of reproduction is Vegetative propagation?

Asexual

what are the natural Vegetative propagation methods used by a plant?

Bulbs

Rhizomes

Stolon’s

Suckers

Tubers

what are Rhizomes and stolons

stem structures that grow horizontally away from parent plant

nodes where new shoots and roots develop

example of plant that uses rhizomes

bamboo

difference between stolon/ runners and rhizomes

stolons → grow above the ground

rhizomes → grow underground

example of plant with stolons/ runners

strawberries

what are suckers ?

shoots that grow from sucker buds on shallow roots of a parent plant

Example of plant with suckers

Elm trees

what are Tubers

underground structure that acts as food stores in plants

covered in eyes → sprouts and form new plant

example of plant with tubers

potatoes

what are bulbs

underground food stores used by plants

new bulbs develop from old bulb → form new individual plants

what are the methods of Artificial Vegetative propagation

Take cuttings

Grafting

layering

Grafting method

join the shoot of a plant to a growing stem and root of another plant

layering method

bending the stem of a growing plant downwards → enters soil and grows into new plant

risk assessment of producing clones from cuttings

be careful with sharp tools

some plants may cause irritation on skin so wear gloves

How to produce clone from cutting ( Image)

How to produce a clone from cutting method

use scalpel to take a cutting from end of stem of parent plant

cutting should have only 1 leaf at tip ( remove rest) ( Reduce transpiration rate)

dip lower end of cutting in rooting powder ( contains hormones that promote root formation) ( Auxins)

place cutting in a pot with growth medium

cutting needs a warm / moist environment → propagator

when cutting has formed roots → plant it elsewhere to continue growing

In clone cutting experiment how to provide cutting with warm environment ?

Cover the pot with plastic bag or put in propagator

what are the artificial and Natural way of cloning a plant

Artificial Vegetative propagation ( man made)

Natural Vegetative propagation

method of using tissue culture to produce clone

stem cells are taken from stem / root tips from parent plant

cells are sterilised then placed on culture medium ( containing glucose + amino acids + plant hormones)

carried out under aseptic conditions → cells divide into a mass of undifferentiated cells

when cells grow into plant → take out of medium and plant in soil

develop into genetically identical plants to parent

method of using tissue culture to produce clone (Image)

Why are cells sterilised in tissue culture

To kill any microorganisms → fungi and bacteria compete with cells for nutrients decreases growth rate

why is tissue culture used?

clone plants that don’t readily reproduce

endangered

rare

grow whole plants from GE plant cells

Uses of micropropagation ?

produce fields full of GE pest-resistant crop

what is micropropagation?

tissue culture is used to mass produce clone plants quickly

Micropropagation ( Image)

Disadvantages of Artificial plant cloning

No genetic variation → can be killed by single disease

expensive due to high energy use

unfavourable genetic characteristics are passed on to clone

tissue culture can be contaminated → loss of plant being cultured.

Advantages of Artificial plant cloning

Produces plants quicker

favourable genetic characteristics are passed on to clone

plants reproduce in any season

requires less space

How are natural clones produced by animals

embryos splits early into development → produces genetically identical offspring ( identical twins)

what are the techniques to artificially clone animals

Artificial embryo twinning

somatic cell nuclear transfer ( SCNT)

Artifical embryo twinning ( Image)

Artificial embryo twinning method

extract egg cell then fertilise in a Petri dish

fertilised egg → divides forming a embryotic cells ( in vitro)

embryotic cells are separated into a petri dish → develops into embryo

embryo are implanted into surrogate mother

embryos develop inside mother → genetically identical offspring are born

similarities between SCNT and artificial embryo twinning

Both involve surrogate mother

both create genetically identical clones

SCNT method

somatic cell is removed from animal A → extract nucleus

An oocyte ( immature egg cell) is taken from animal B → remove nucleus to form an enucleated oocyte

insert nucleus from A into enucleated oocyte → they fuse and divide → embryo

embryo implanted to surrogate mother.

what are some of the uses of animal cloning

Research purposes → test new drugs

Agriculture → increase number of animals with favourable characteristic

Save endangered animals from extinction

SCNT ( Image)

How can scientists use cloning to replace damaged tissues

Clone embryonic stem cells → totipotent

replace damaged tissues

Genetically identical to patient → don’t get rejected by immune system

Advantages of animal cloning

Develop new disease treatment → less chronically ill people

Favourable genetic characteristic is passed on → cow with lots of milk → calves lots of milk

increase population of endangered species → preserve biodiversity

Animals can be cloned at any time → dont have to wait till breeding season

disadvantages of animal cloning

Time-consuming and expensive

no genetic diversity → all susceptible to same disease

unethical → clones live shorter then natural offspring and involves destroying embryos

What is biotechnology ?

industrial use of living organisms to produce food, drugs and other products

why are micro-organisms mainly used in biotech ?

can be grown any time of the year

Can be grown on inexpensive material → economical to use

Ideal growth conditions are easily produced

short life cycle → grow quickly

what are the organisms used in biotechnology

microorganisms → fungi and bacteria

enzymes

what are the different types of enzymes used in biotech

Intracellular enzymes → enzymes are contained within cells

extracellular enzymes → enzymes are naturally secreted

isolated → enzymes that aren’t contained within cells

Disadvantages of intracellular enzymes

Expensive to extract the enzymes

what are the food industries / prodcution that use microorganisms

Baking

Brewing ( making beer)

Cheese making

Yoghurt production

Brewing ( making beer) process biotech

yeast ( saccharomyces cerevisiae) added to grain

yeast anaerobically respires using glucose from grain ( produces CO₂ + ethanol)

Baking process biotech

Yeast alcoholic fermentation → CO₂ makes bread rise

cheese making process biotech

Rennet’s contain the enzyme chymosin which clots the milk

chymosin can be obtained from GM yeast cells to produce enzymes

Lactic acid bacteria convert lactose in milk → lactic acid

helps cheese solidify

Yoghurt production

lactic acid bacteria → clots the milk and causes it to thicken

what are the uses of micro-organisms in medicine production

Penicillin production

insulin production

how is penicillin produced in biotech

the fungus (Penicillium chrysogenum) grown under stress in industrial fermenters

produce the antibiotic penicillin → kills bacteria, stops them competing for resources

penicillin produced is collected and processed

which products use industrial fermenters

Insulin production

Penicillin production

how is Insulin produced in Biotech

made by GM bacteria with gene for human insulin inserted in their DNA

bacteria is grown in industrial fermenter → insulin produced is collected and purified

What is bioremediation ?

Process of using organisms to remove pollutants from contaminated sites

Break down pollutants into less harmful products

How is bioremediation used naturally ?

pollutant- removing bacteria are given extra nutrients and enhanced growing conditions to allow maximum multiplication.

Example of microorganism used to make single-celled protein

The fungus Fusarium venenatum used to make Quorn

Advantages of using micro-organisms in food production

Micro-organisms can be grown quickly and easily → inexpensive

microorganisms can be cultured anywhere → used to tackle malnutrition in developing countries

single-cell protein is a healthier alternative to animal protein

more ethical → doesn’t involve animals dying

disadvantages of using micro-organisms in food production

single-cell protein has different flavour and texture to animal meat

easily contaminated by unwanted bacteria → conditions are ideal for other micro-organisms

people wont eat food grown using waste products

single- cell protein is dangerous when consumed in high amounts

What is meant by a culture ?

Population of one type of microorganism grown under controlled conditions

where are cultures grown

fermentation vessels

what are the methods of culturing micro-organisms

Batch fermentation → grown in individual batches in fermentation vessel

Continuous fermentation → microorganism are continually grow.

differences between batch and continuous fermentation

Batch → closed culture, waste products aren’t removed and extra nutrients are added

Continuous → Open culture, nutrients are added and waste products are removed at a constant rate

Fermentation vessel ( Image)

control variables in fermentation vessel

pH

Temperature

O₂ supply

Nutrients concentration

contamination

Function of pH probe in fermentation vessel

monitors pH → optimum pH → highest product yield as optimum enzyme activity

fastest rate of reaction

Function of water jacket in fermentation vessel

keeps temperature at optimum → highest product yield as optimum enzyme activity

fastest rate of reaction

Function of sterile air in fermentation vessel

keeps the volume of O₂ at optimum for respiration → increases product yield as microorganisms can respire to provide energy for growth

Function of paddles in fermentation vessel

Stir the culture ensures mixing of nutrients/ O₂→ increases product yield as microorganisms have constant access to nutrients needed for growth.

why are fermentation vessels sterilised?

sterilised with superheated steam to kill any unwanted microorganisms → stops contamination

increases product yield as microorganisms aren’t competing with other organisms

What is a closed culture

Growth occurs in a vessel that is isolated from the external environment → extra nutrients aren’t added and waste products aren`t removed

Examples of a close culture

Batch fermentation

What are the four phases of a standard growth curve

Lag phase

Log phase

Stationary phase

Decline phase

Standard growth Curve

Growth curve for continuous fermentation ( Image)

Growth curve continuous fermentation features

No stationary and decline phase → Open culture

nutrients are constantly being added and waste is constantly being removed

waste can`t build up to toxic levels

closed culture growth curve ( Lag phase)

population size increases slowly → microorganism have to make enzymes and other molecules before they reproduce

reproduction rate is low

closed culture growth curve ( Log phase)

Population size increases quickly → favourable culture conditions for reproduction ( lots of food and little competition)

number of microorganisms more than doubles at regular intervals

closed culture growth curve ( stationary phase)

population size stay level → micro-organism death rate = reproductive rate

micro-organisms die → toxic waste products build up and lack of food

closed culture growth curve ( decline phase)

population size decreases → death rate is greater then reproductive rate

toxic waste products increases + food is limiting

equation to work out cell population

what does Log show

How many times a 10 has been x 10

Log values (image)

How to work out rate of growth during time period on micro-organism time graph

work out gradient between the 2 time periods

How to convert log number of cells into actual number of cells

10ⁿ

n = log number of cells

how to culture micro-organism in lab

Use a Agar plate (sterile Petri dish)

how to improve growing conditions when culturing micro-organisms

Add nutrients

what are the ways to culture microogranisms

In a Agar plate

in a Broth

how are micro-organisms provided in when culturing

In a liquid broth ( distilled water + nutrients)

Differences between culturing micro-organism in a broth and Agar plate

Agar plate → culture medium is solid

Broth → culture medium is liquid

Advantages using broth and Agar plate when culturing

Agar plate → allows isolation and separation of microorganism → easier to study individual colony

Broth → Supports fast microorganism growth in large volumes

Apparatus used when culturing micro-organisms

pipette

incubator → micro-organism can grow

glass spreader ( sterile) → spreads on whole surface of Agar

wire inoculation loop → transfer sample on the plate

what are Aseptic techniques and function

Prevent contamination of culture by unwanted micro-organisms

unwanted microorganisms → health risks , affect growth of micro-organism cultured, experiment gives in accurate results

Examples of aseptic techniques when culturing

sterilise all apparatus used after and before transfer of culture ( heating with flame for 5 seconds)

minimise the time Agar plate is open, put lid on ASAP → prevents airborne microorganisms from contaminating culture

Pass broth container through flame after opened and closed → causes air to move out → preventing unwanted organisms from falling in

Control variables when culturing

Temperature

pH

O₂ supply

nutrients levels

Why are Agar plates incubated inverted in culturing?

Stops any condensation formed on the lid from entering the agar

What to do if colonies overlap and you cant count them when culturing

Serial dilution of micro-organism on broth

plate on agar