A2 LEVEL

Chapter 19
Genetic Technology
Chapter Outline
Technology Function Application
1. Polymerase Chain Clone and amplify DNA Too many…
Reaction (PCR)
• Role of Taq polymerase

Updated on 12/8/21 by Beh SJ @behlogy

 Chapter Outline
Technology Function Application
2. Recombinant DNA To make many copies of Large-scale production of human
technology / Gene Cloning a gene or protein proteins as drugs
E.g. insulin, factor VIII for haemophilia,
• Properties of plasmids adenosine deaminase for SCID
• Promoters
• Marker genes (e.g. Genetic engineering of
fluorescent genes / genes crop plants and livestock
coding for easily stained
substances) • GM salmon
• Role of restriction • Insect resistance
endonucleases, reverse E.g. Bt maize, Bt cotton
transcriptase and ligase • Herbicide resistance
E.g. maize, cotton, tobacco, oil
seed rape

• Ethical and social implications

of using GMOs in food
production

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Chapter Outline
Technology Function Application
3. Gel electrophoresis To separate fragments of Genetic fingerprinting /
• For protein and for DNA DNA according to length DNA profiling
• Probes OR • Paternity testing
To separate diff proteins • Criminal Investigations
according to
mass/charge Genetic screening
• Breast cancer (BRCA1, BRCA2)
Distinguish btwn alleles • Genes for haemophilia, SCA,
of a gene Huntington’s Disease and CF

Gene Therapy
• Vectors (viruses, liposomes,
naked DNA)
• SCID, inherited eye disease, CF
• Social and ethical
considerations

Updated on 12/8/21 by Beh SJ @behlogy

 Chapter Outline
Technology Function Application
4. Microarrays Identify expressed genes • Genetic screening
by detecting mRNA • Drug testing
OR • Identify genes that are
Distinguish btwn alleles switched on/off in diseases
of a gene

5. Bioinformatics and DNA Collection, processing • Role in drug development

sequencing and analysis of biological E.g. against Plasmodium
(Extra! Not in syllabus) info and data using
computer software • Protein and DNA seq provide
evidence for evolution
Allows comparison of
DNA and protein
sequences
6. CRIPSR-Cas9 Cut DNA at specific loci Cure viral diseases and genetic
(Extra! Not in syllabus) and allow genome diseases, even cancer!
editing

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Technology 1
POLYMERASE CHAIN REACTION (PCR)
Function: Clone and amplify DNA
Updated on 12/8/21 by Beh SJ @behlogy
Chapter Outline
Technology Function Application
1. Polymerase Chain Clone and amplify DNA Too many…
Reaction (PCR)
• Role of Taq polymerase

Updated on 12/8/21 by Beh SJ @behlogy

 Polymerase Chain Reaction (PCR)
• Rapid and efficient process
• Function: clone and amplify DNA
• Amplify = production of many copies of a length of DNA
• Only small sample of DNA needed

Components needed:
1. Template DNA
2. Taq polymerase
• Heat-stable DNA pol from Thermus aquaticus bacteria
3. Buffer
• Contains KCl and MgCl2
4. 4 types of nucleotides (A, T, C, G)
5. 2 Primers
• Short sequence of ~20bp of
single-stranded RNA/DNA
• Complementary base pair to start
and end of target region for amplification
Updated on 12/8/21 by Beh SJ @behlogy
 Polymerase Chain Reaction (PCR)
1. Denaturation (95⁰C)
• DNA strands separate / denature into 2
strands by heat
• Hydrogen bonds between DNA strands breaks
• Bases are exposed
• Produce template strands for copying

Updated on 12/8/21 by Beh SJ @behlogy

 Polymerase Chain Reaction (PCR)
2. Annealing (60-65 ⁰C)
• Primer added
• Primers anneal / bind to specific section of DNA
• Via complementary base pairing
• New hydrogen bonds form

Role of primers:
• Bind to target region for amplification
• Acts as a starting point for for Taq polymerase to bind
→Taq polymerase only binds to double-stranded DNA
and add new nucleotides to an existing strand
• Reduce reannealing of separated strands

Updated on 12/8/21 by Beh SJ @behlogy

 Polymerase Chain Reaction (PCR)
3. Extension (70-75 ⁰C)
• Taq polymerase binds to primer
• Synthesises new DNA strands
• Complementary to the DNA template strands

• Taq polymerase has a high optimal temperature

/ is heat stable
• Does not need replacing each cycle

Updated on 12/8/21 by Beh SJ @behlogy

 Polymerase Chain Reaction (PCR)
• DNA is heated again to separate strands
• The process is repeated until sufficient DNA is produced (usually ~30 cycles)
• The number of DNA molecules doubles every cycle = efficient process!
• Number of DNA double helix copies made from
one starting molecule after n cycles of PCR = 2n

PCR simulation = http://learn.genetics.utah.edu/content/labs/pcr/

Currently not available  Watch it here = https://www.youtube.com/watch?v=0jlWKw5qEP8
Updated on 12/8/21 by Beh SJ @behlogy
 Polymerase Chain Reaction (PCR)
Advantages:
• Rapid, efficient
• Only small sample of DNA needed for amplification of DNA
• Process is automated in a thermal cycler

Disadvantages:
• Need to know the precise DNA sequence beforehand to design primers
• DNA amplification have to be in shorter fragments than gene cloning in
bacteria (Genetic Technique 2)

Updated on 12/8/21 by Beh SJ @behlogy

Applications of Polymerase Chain Reaction (PCR)
Function: clone and amplify DNA

Applications:
• DNA sequencing
→ Able to amplify small amount of DNA extracted for sequencing
(E.g. in fossils, blood sample)
• DNA profiling
→ Able to amplify small amounts of DNA extracted (E.g. at crime scene)
• Recombinant DNA Technology
→ Amplify DNA/gene needed for insertion into plasmid
• Genetic screening
→ To identify mutations/disease genes/DNA from pathogens
→ Use primers complementary to target gene
→ To identify and amplify target gene in specific
→ Gel electrophoresis used to isolate gene
Updated on 12/8/21 by Beh SJ @behlogy
Genetic Technology 2
RECOMBINANT DNA TECHNOLOGY /
GENE CLONING
Function: to make many copies of one gene / protein
Updated on 12/8/21 by Beh SJ @behlogy
Chapter Outline
Technology Function Application
2. Recombinant DNA To make many copies of Large-scale production of human
technology / Gene Cloning a gene or protein proteins as drugs
E.g. insulin, factor VIII for haemophilia,
• Properties of plasmids adenosine deaminase for SCID
• Promoters
• Marker genes (e.g. Genetic engineering of
fluorescent genes / genes crop plants and livestock
coding for easily stained
substances) • GM salmon
• Role of restriction • Insect resistance
endonucleases, reverse E.g. Bt maize, Bt cotton
transcriptase and ligase • Herbicide resistance
E.g. maize, cotton, tobacco, oil
seed rape

• Ethical and social implications

of using GMOs in food
production

Updated on 12/8/21 by Beh SJ @behlogy

Recombinant DNA Technology

1. Identify and isolate mRNA /

DNA from organism

2. Cut target gene and

plasmid DNA

6. Grow bacteria in
3. Recombine/join gene fermenters for
to plasmid DNA large-scale production

5. Identify modified /
transformed bacteria

4. Insert recombinant
plasmid into bacteria
Updated on 12/8/21 by Beh SJ @behlogy
E.g. Production of human insulin
1. Obtain mRNA for human insulin
• From β cells of islets of Langerhans of pancreas
• Reverse transcriptase to make cDNA
(single-stranded complementary DNA)
from mRNA
• DNA polymerase used to make
double-stranded cDNA from ssDNA
• Final result: gene with no introns, shorter DNA

• DNA may be amplified using

Polymerase Chain Reaction (PCR)

Updated on 12/8/21 by Beh SJ @behlogy

 2. Cut target gene and plasmid DNA
• Use restriction enzyme to cut the gene
• Restriction sites should be present at both sides of gene

• Obtain plasmids from bacteria

• Cut plasmid at 1 restriction site using SAME restriction enzyme
• Complementary sticky ends produced

Updated on 12/8/21 by Beh SJ @behlogy

 Restriction Enzymes / Endonucleases
• From bacteria
• Recognize, bind and cut DNA at a specific sequence, called a restriction site
• Hydrolyses/cleaves the phosphodiester bond btwn nucleotides
• Diff kind of RE cuts at a diff specific seq

• Most restriction sites are palindromic sequences

• Restriction enzymes produces sticky ends or blunt ends

At the ‘sticky ends’:

• Result of a staggered cut
• A few unpaired nucleotides at ends
• Able to easily form H bonds /
complementary base pair

At ‘blunt ends’
• NO unpaired nucleotides
Updated on 12/8/21 by Beh SJ @behlogy
 Plasmids
• Small, circular, double-stranded DNA
• Found in bacteria, but now usually artificially made in lab

Frequently used in gene technology because:

• Small so can be inserted into cells
• Circular so more stable / not damaged by host cell enzymes
• Plasmids act as vector to deliver desired genes to bacteria
• Easy to extract from bacteria
• Can be taken up by bacteria due to low molecular mass/small
• Has high copy number
– many copies can be present in one bacterial host cell
• Replicate independently within bacteria
– able to clone/replicate any genes inserted into them
• Has specific DNA sequences needed!

Updated on 12/8/21 by Beh SJ @behlogy

 DNA Sequence Function
Origin of • Allows bacterial DNA polymerase to bind and replication to be
replication initiated
• For restriction enzyme to cut and produce sticky ends, so gene of
Restriction site interest can be inserted
• Can have multiple to be cut by diff RE → has multiple cloning sites
• Initiates transcription
• Allows binding of RNA polymerase /
• transcription factors
• Ensures correct strand is used as template
Promoter • Diff promoters determine diff level of
expression and where it is expressed
• E.g. use promoter upstream of lacZ gene
in lac operon – target gene will be
expressed in the presence of lactose
• Gene of interest is inserted close to / into marker gene
• Both the target gene and marker gene are expressed
• Helps to recognise recombinant plasmids / modified bacteria /
Marker Gene
transgenic organisms
• E.g. genes for antibiotic resistance, genes for fluorescent / easily

Plasmids stained substances

Updated on 12/8/21 by Beh SJ @behlogy
Restriction Enzymes

Updated on 12/8/21 by Beh SJ @behlogy

 3. Recombine/join gene to plasmid DNA
• Mix cDNA / insulin gene with plasmid
• DNA ligase seals nicks in sugar-phosphate backbone
→ Catalyses the formation of phosphodiester bond
• To form recombinant DNA

Updated on 12/8/21 by Beh SJ @behlogy

4. Insert recombinant plasmid into bacteria
1. Recombinant plasmids mixed with bacteria
2. Treat bacteria with a solution of Ca2+ ions and allow to cool
3. Apply heat shock (~42oC) or use electroporation to increase chances of
plasmids passing through cell surface membrane

Only ~1% bacteria will take up recombinant plasmids = transformed

Updated on 12/8/21 by Beh SJ @behlogy

5. Identify modified / transformed bacteria
• Marker genes in the plasmid helps to recognise recombinant plasmids /
modified bacteria / transgenic organisms
• E.g genes for antibiotic resistance, genes for fluorescent /
easily stained substances
• Gene of interest is inserted close to OR into marker gene
• Both target gene and marker gene are expressed
Gene of interest is inserted into marker gene
Marker gene is disrupted!
Gene of interest is inserted close to marker gene There is another marker gene that is expressed together
Downstream of same promoter Extra function: easy to identify recombinant plasmid

Updated on 12/8/21 by Beh SJ @behlogy

5. Identify modified / transformed bacteria
Ways things can go wrong:
1. Bacteria did not take up recombinant plasmid
2. Gene of interest did not join with plasmid…
Cut plasmid just rejoined with itself

Ways to identify transformed bacteria:

a) Use antibiotic selection
b) Use green fluorescent protein (GFP)
c) Use an easily stained substance (β galactosidase, GUS)

Updated on 12/8/21 by Beh SJ @behlogy

 a) Antibiotic Selection
• Genes for antibiotic resistance used as markers
E.g. ampicillin resistance gene (ampr) , tetracycline resistance gene (tetr)

• Grow on agar containing the antibiotic (ampicillin)

• Bacteria with plasmid will be able to survive
• Bacteria without the plasmid dies

• Then make a replica plate by using sponge/velvet pad

• Grow bacteria on agar containing 2nd antibiotic (tetracycline)
• Bacteria with recombinant plasmid will die

Updated on 12/8/21 by Beh SJ @behlogy

 a) Antibiotic Selection
BUT PROBLEM!
• Risk of spread of antibiotic resistance to other bacteria of same / diff species
• Plasmids are easily transferred between bacteria via conjugation
• This makes the use of antibiotics less effective in disease-causing bacteria
• Slower process for identification of transformed bacteria as well
→ So… use other methods to identify modified bacteria

Updated on 12/8/21 by Beh SJ @behlogy

 b) Use Green Fluorescent Protein (GFP)
• Gene for fluorescent substances used as marker
E.g. gene for Green fluorescent protein (GFP)

• From jellyfish
• GFP emits bright green light
• When exposed to UV

• Bacteria with plasmid will express GFP and will

appear green

Updated on 12/8/21 by Beh SJ @behlogy

 c) Use an easily stained substance
• Use gene that codes for easily stained substances as marker
E.g. gene for β galactosidase aka lacZ gene
E.g. gene for GUS enzyme

• Enzyme splits a special non-blue substrate into product that is blue

• Bacteria with plasmid will become blue

Updated on 12/8/21 by Beh SJ @behlogy

 Advantages of using genes for fluorescent /
easily stained substances as markers
• Avoid use of antibiotics
• More economical / time saving / labour saving
• Visible colour is easy to identify / detect
• Enable identification of transformed cells AND transgenic organisms
• No known risk / ill effect on GM organism

Updated on 12/8/21 by Beh SJ @behlogy

 6. Grow bacteria in fermenters for
large-scale production
• Allow transformed bacteria cells to multiply/clone
• Grow in large-scale culture / fermenter
• E. coli can divide once every 20mins

• Bacteria produces multiple copies of the gene /

protein product (in this case, human insulin)
• Bacteria has replication, transcription and
translation machinery to copy the gene and express
the protein product
(e.g. DNA polymerase, RNA polymerase, ribosomes)

• Insulin extracted and purified to be sold on market

Updated on 12/8/21 by Beh SJ @behlogy

 Advantages of Recombinant DNA Technology
• Conventional method = extract insulin from pancreas of pigs or cattle

Advantages of producing human insulin by gene technology:

1. Chemically identical to human insulin
→ exact fit to insulin receptors on target organs
→ does not stimulate the immune system
→ faster response
→ fewer side affects
→ less / no risk of allergic reaction
2. Effective in people who have developed tolerance to animal-derived insulin
3. Avoid ethical issues related to religion & use of animal products
→ no killing of animals
4. Lower cost of purification and processing
5. Mass production = large and constant reliable supply all year round
6. Less risk of contamination/infection
→ no risk of transfer of disease
7. Potential to engineer / improve recombinant proteins
Updated on 12/8/21 by Beh SJ @behlogy
 Applications of Recombinant DNA Technology
• Production of pharmaceuticals
→ No modifying of protein in bacteria (bcs no membrane-bound organelles)
→ Can genetically modify eukaryotic cells to produce human proteins
E.g. yeast cells, insect larvae cells, mammalian cells

Human Protein Treatment for Produced in

Human insulin Diabetes Recombinant bacteria
Helps blood clot in haemophilia GM hamster kidney and ovary
Factor VIII
patients cells (in fermenter)
Help in development of B & T cells in
Adenosine GM insect larva of the cabbage
severe combined immunodeficiency
deaminase (ADA) looper moth
disorder (SCID) patients
Antithrombin Stop blood coagulation Milk of GM goats
Alpha-antitrypsin Emphysema Milk of GM sheep

• Used in genetic engineering to produce GMOs in agriculture

Updated on 12/8/21 by Beh SJ @behlogy

 Genetic Engineering
• Manipulation of genetic material to modify specific characteristics of an organism
• Extraction of genes from one organism / synthesis of genes
• In order to place them in another organism
• Of the same or diff species

New hosts:
• Have recombinant DNA (rDNA) = combination of DNA from two or more sources
• Expresses the new gene product / protein

• Genetically modified (GM) /

transgenic organisms
= organisms with any altered DNA

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Editing
• Form of genetic engineering
• Involving the insertion, deletion or replacement of DNA
• At specific sites in the genome

Updated on 12/8/21 by Beh SJ @behlogy

 GMOs in Agriculture
Aims of using genetic engineering to produce GMOs in agriculture:
• improve quality of crop plants and livestock
• Increase yield of crop plants and livestock
→ Solve world food demand

Updated on 12/8/21 by Beh SJ @behlogy

 GMOs in Agriculture
Livestock can be engineered to:
• Have high growth rate
• Grow larger
• Have higher yield (e.g. milk, meat)

Crop plants can be engineered to:

• Have higher yield
• Better quality / taste
• Delayed ripening of fruits (to increase shelf life)
• Additional nutritional benefits
• Resist disease/pests/insects (so less pesticides used)
• Resist herbicides (to reduce competition from weeds)
• Grow in adverse conditions / more tolerant to climate
change (e.g. high salinity, dry, hot, cold climates)
• Grow in poor quality land, require less fertilizer
→ More cost effective / have health benefits
Flavr Savr Tomatoes
→ Less effect on food chain / pollinators
Updated on 12/8/21 by Beh SJ @behlogy
Updated on 12/8/21 by Beh SJ @behlogy
 GMOs in Agriculture
Why use GMOs instead of artificial selection?
• Much faster results
• Allows the retention of other desirable characteristics of
best varieties of species
• Able to use best genes from other species of plants and
even non-plants

But…
• Complicated process
• Expensive
• Not always successful

Updated on 12/8/21 by Beh SJ @behlogy

 GMOs in Agriculture
Important examples:
• GM salmon
• Vitamin A enhanced rice (Golden riceTM)
• Insect resistance
E.g. Bt maize, Bt cotton
• Herbicide resistance
E.g. maize, cotton, tobacco, oil seed rape

Ethical and social implications of using GMOs in

food production

Updated on 12/8/21 by Beh SJ @behlogy

 GM Atlantic Salmon
• AquAdvantage® salmon
• Grows to market weight in about 16-18 months
• Normal salmon needs 32-36 months

• Genes for growth hormone regulator

transferred from diff species of salmon
• Genes for promoter from another diff species

Benefits:
• High yield
• Consistent yield all-year round
• Conserve wild fish populations
• All modified salmon eggs are triploid and sterile
→ so impossible for them to breed among themselves and with other salmon
→ eliminate impact to wild population
Updated on 12/8/21 by Beh SJ @behlogy
 Insect-Resistant Crops
Bt Crops
E.g. Bt cotton, Bt maize
• Bt cotton protected against boll weevils

• Bt maize protected against corn borer caterpillars

• Corn borer eats leaves and burrows into stalk
→ plant cannot support the ears of corn

• Gene for Bt toxin from bacteria Bacillus

thuringiensis inserted into maize / cotton
• Lethal to insects but not other animals
• Crop able to produce own insecticide

Updated on 12/8/21 by Beh SJ @behlogy

Insect-Resistant Crops
Bt Crops
Benefits of Bt Crops:
• Increased yields

• Only kills specific insects that eats it and

does not kill beneficial insects
E.g. pollinators / bees / predators of pests
→ Conserve biodiversity / food web

• Less pesticide used

→ Reduce risk of pesticide affecting other non-target species in the same
environment
• Less risk of harm to humans from spray drift / pesticide residues on food

Updated on 12/8/21 by Beh SJ @behlogy

Herbicide-Resistant Crops
Glyphosate-resistant crops
E.g. tobacco, oil seed rape, Roundup Ready soybean
• Genetically modified to be resistant to herbicides containing
glyphosate (e.g. Roundup)
• Glyphosate inhibits enzymes involved in amino acid synthesis

• Gene from bacterium Agrobacterium coding for enzymes with same

function that are not affected by glyphosate
• Herbicide resistant gene acts as marker gene
→ Herbicides will have no effect on plant, only the weeds

Advantages:
• Can control/kill weeds
→ reduce competition
→ increase yield
• Less manual weeding needed
Updated on 12/8/21 by Beh SJ @behlogy
 Herbicide-Resistant Crops
Glyphosate-resistant crops
Disadvantages:
Environmental
1. Intensive use of herbicides
→ High toxicity of more herbicide left after use
→ harmful to humans

2. Cross-pollination with wild plants / organic crops

→ Result in new more resistant weeds = “superweeds”
→ Superweeds are selected for by herbicide use
→ Outcompete and lose traditional varieties of plants
→ Loss of genetic diversity
→ Cause effect on rest of food chain = less food or shelter for other species
https://www.thestar.com.my/news/nation/2018/01/16/farmers-go-against-the-grain-weed-interbreeding-sees-clearfields-future-grow-cloudy/

Solution: Use sterile seeds/plants to avoid superweeds

Updated on 12/8/21 by Beh SJ @behlogy
 Herbicide-Resistant Crops
Glyphosate-resistant crops
Disadvantages:
Economic
• High cost of GM seeds
→ Seeds are sterile and cannot use own seed
→ Too expensive for people to buy / farmers to sell
→ Difficult for farmers in developing countries to obtain GM seed
→ Might reduce efforts to relieve poverty
→ Under-developed countries become more dependent on other counties

• Cost of herbicide
→ cost of problems with pollution
→ cost of human health problems
→ loss due to contaminated crops of organic farms

Updated on 12/8/21 by Beh SJ @behlogy

Other ethical and social implications of
using GMOs in food production
• GM production cost is expensive
→ Does it outweigh benefits? It has only 1% success…

• Monopoly / too much power held by multinational companies

• People may avoid / refuse to buy for GM food

• No long-term studies done on effects on human health

• Possible allergic reactions in humans / adverse effects on the
immune system

https://www.youtube.com/watch?v=7TmcXYp8xu4

Updated on 12/8/21 by Beh SJ @behlogy

Updated on 12/8/21 by Beh SJ @behlogy
https://gmoanswers.com/why-do-some-people-say-or-think-gmos-are-bad

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Technology 3
GEL ELECTROPHORESIS
Function: To separate fragments of DNA according to length OR
To separate diff proteins according to mass/charge
Updated on 12/8/21 by Beh SJ @behlogy
 P5
Chapter Outline
Technology Function Application
3. Gel electrophoresis To separate fragments of Genetic fingerprinting /
• For protein and for DNA DNA according to length DNA profiling
• Probes OR • Paternity testing
To separate diff proteins • Criminal Investigations
according to
mass/charge Genetic screening
• Breast cancer (BRCA1, BRCA2)
Distinguish btwn alleles • Genes for haemophilia, SCA,
of a gene Huntington’s Disease and CF

Gene Therapy
• Vectors (viruses, liposomes,
naked DNA)
• SCID, inherited eye disease, CF
• Social and ethical
considerations

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis
Components Functions
Diff gel, diff impedance
Agarose gel (DNA) or
Even spaces between the gel molecules so DNA / protein can
Polyacrylamide gel
move within gel properly
(proteins) with wells
Wells to load DNA / protein
Electrodes + electric field Molecules with net negative charge will move towards
positive electrode
Buffer solution Maintain constant pH

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis
Components Functions
Visible dye to track DNA migration
Loading dye → Ensure electrophoresis is stopped before DNA / protein runs
off the gel
Dye that stains the colourless DNA / protein
Staining agent
E.g. Ethidium bromide causes DNA to fluoresce under UV light
DNA ladder or Has DNA fragments or substances of known lengths / mass
protein ladder → Can estimate the length / mass of samples by comparing them
to the ladder
Gene probes (DNA) To locate a specific DNA sequence

Loading dye Protein ladder

DNA ladder

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis of DNA
Procedure:
1. Obtain sufficient quantity of DNA
• DNA can amplified using PCR or gene cloning in
bacteria

2. Cut DNA into fragments using restriction enzymes

• Many DNA fragments of diff lengths produced

3. DNA samples are mixed with loading dye and

staining agent

4. Gel is covered with buffer solution

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis of DNA
5. DNA fragments are loaded into wells at the negative end of the gel (cathode)
• DNA ladder with DNA fragments of known lengths is usually run through the
gel at the same time

6. A direct current is applied through the gel

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis of DNA
7 . DNA is attracted towards the positive electrode (anode)
• Separation due to electric field / potential difference
• Bcs DNA is negatively charged due to the phosphate groups

• Gel acts a molecular sieve

• Shorter fragments of DNA move faster and
further per unit time through the gel than
larger fragments
• Due to less impedance of gel
• DNA fragments separate and arranged
in order of size

Updated on 12/8/21 by Beh SJ @behlogy

 P5

Gel Electrophoresis of DNA

8. Visualise bands under UV light
• Staining agent causes DNA to fluoresce
• Each band is millions of DNA pieces
of the same size

• Compare position with reference DNA

to estimate length of fragments

Simulation = http://learn.genetics.utah.edu/content/labs/gel/
Updated on 12/8/21 by Beh SJ @behlogy
 P5
Gel Electrophoresis of DNA
9. DNA pieces transferred to membrane / nylon / nitrocellulose / absorbent paper
• This is called Southern blotting

10. DNA heated to separate strands

Updated on 12/8/21 by Beh SJ @behlogy

 P5
Gel Electrophoresis of DNA
11. DNA cooled and gene probes are added
• Gene probes are used to locate a specific DNA sequence or gene
• Gene probes = short, single stranded DNA (ssDNA)
• ‘Labelled’ / attached with radioactive / fluorescent substance
• Can complementary base pair with specific DNA fragments

12. View DNA fragments under UV light / X-ray film

Updated on 12/8/21 by Beh SJ @behlogy

 Applications of DNA Gel Electrophoresis
• Can use to distinguish between the alleles of a gene

• Genetic fingerprinting / DNA profiling

• Paternity testing
• Criminal Investigations

• Genetic screening
• Breast cancer (BRCA1, BRCA2)
• Genes for haemophilia, SCA, Huntington’s Disease and CF

• Gene Therapy
• Vectors (viruses, liposomes, naked DNA)
• SCID, inherited eye disease, CF
• Social and ethical considerations

Updated on 12/8/21 by Beh SJ @behlogy

 Genetic Screening
• Aka genetic testing
• Analysis of a person’s DNA to check for presence of a particular allele
• DNA obtained from tissue samples (e.g. blood)
• Can be carried out on embryo, fetus, newborn or adults

• Important examples: breast cancer, haemophilia, sickle cell anaemia,

Huntington’s disease, cystic fibrosis (CF), Down syndrome

Updated on 12/8/21 by Beh SJ @behlogy

 Genetic Screening
Type of Genetic Screening Who is it for?
1. Pre-implantation genetic Newly formed embryo from IVF
diagnosis (PGD) Before implantation into uterus of mother
2. Prenatal screening Unborn children / fetus
3. Newborn screening Newborn babies
Individuals with a family history of a genetic
4. Carrier screening condition / cancer
Potential parents

Updated on 12/8/21 by Beh SJ @behlogy

In vitro fertilisation (IVF) Chap 18 Recap
1. Hormone treatment used to induce superovulation
2. Many oocytes harvested from female
3. Obtain fresh / frozen sperm
4. Oocytes mixed with sperm
– Ideally genetically different
5. Conduct genetic test and select
embryos that are most likely to
survive
6. Embryo transfer!
– Embryo implanted in uterus of donor female
– OR may use similar species / non-rare breed as surrogate mother
– OR freeze embryos and store for long time in frozen zoo

E.g. IVF + Frozen Zoos: https://www.youtube.com/watch?v=0UsoDqECYb8

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Screening

7.

6. Pre-implantation
genetic diagnosis (PGD)
and preselection of
embryos

Updated on 12/8/21 by Beh SJ @behlogy

1. Pre-implantation genetic diagnosis (PGD)
• Newly formed embryo from in vitro fertilisation (IVF)
• Tested before implantation into uterus of mother

Procedure
1) Embryo biopsy = removing a cell from an embryo
• At 8-cell stage (day 4/5)

2) PCR to amplify DNA

3) Gel electrophoresis
• Analyse DNA for faulty alleles to determine if embryo has genetic disease
• E.g. haemophilia, Huntington’s disease, cystic fibrosis

4) Pre-selection = select only embryo without faulty alleles for implantation

Updated on 12/8/21 by Beh SJ @behlogy

1. Pre-implantation genetic diagnosis (PGD)
Social or ethical implications:
• Identify whether embryos from IVF have a genetic condition
→ Avoid implantation of embryos with faulty alleles
→ Allows couples to have children who would otherwise choose not to

BUT…
**It’s controversial!**

Ethical concerns:
• Embryos might be destroyed if not pre-selected for implantation
• Could lead to selection based on gender or specific traits (“designer babies”)
• Contrary to beliefs / values
• Genetic disease may not develop

Updated on 12/8/21 by Beh SJ @behlogy

 2. Prenatal screening
Methods to obtain tissue samples:
(1) amniocentesis
https://www.webmd.com/baby/video/amniocentesis
(2) chorionic villus sampling (CVS)
https://www.mayoclinic.org/tests-procedures/chorionic-villus-sampling/multimedia/chorionic-villus-sampling-video/vid-20121056

Updated on 12/8/21 by Beh SJ @behlogy

 2. Prenatal screening
Advantages
• Helps to provide early diagnosis for fetuses in utero
→ So can give early treatment when born
→ Allows parents to prepare for the birth of a child who will need treatment for
a considerable time or even throughout life
→ Women can avoid late therapeutic abortions = terminate pregnancy for
medical reasons

Updated on 12/8/21 by Beh SJ @behlogy

3. Newborn screening / 4. Carrier Screening
E.g. haemophilia, sickle cell anaemia, cystic fibrosis (CF),
Huntington’s disease, Down syndrome

E.g. breast cancer

• Faulty alleles of the BRCA1 and BRCA2 genes
• Increase the chance of developing breast cancer
• High risk

If tested positive for the faulty alleles:

• Increased testing to detect cancer early
• Elective mastectomy = removal of breast(s)
before the occurrence / diagnosis of cancer

Updated on 12/8/21 by Beh SJ @behlogy

3. Newborn screening / 4. Carrier Screening
Advantages
• Can identify carriers of genetic conditions
→ Allows couples who are both carriers of a
genetic condition to choose not to have children

• Helps to provide early diagnosis

e.g. Huntington’s disease (symptoms only occur later in life)
→ can prepare for the future:
1. if present, enables lifestyle change / early treatment / regular check-ups
2. preventative treatment may be cheaper than treating disease itself
→ if tested negative, genetic screening removes anxiety

Updated on 12/8/21 by Beh SJ @behlogy

3. Newborn screening / 4. Carrier Screening
Disadvantages
• Test is expensive
• Many mutations still unknown so may still
not lead to positive diagnosis

If person tested positive:

• For some diseases, no treatment is possible
• May lead to anxiety
• May experience social / financial
discrimination (e.g. life insurance refusal)
• May still not develop diseases even though
mutation is present
• Couple may decide to not have children
OR couple may be tested after they have
children
https://www.vox.com/recode/2019/12/13/20978024/genetic-testing-dna-
consequences-23andme-ancestry
Updated on 12/8/21 by Beh SJ @behlogy
 Genetic Counselling
Couple are referred to genetic counsellors if they:
• both have genetic disease (in family) or are carriers
• have history of recurrent miscarriages
• female is an older woman

Genetic counsellor can:

• run pedigree analysis + genetic screening
• explain results of tests / estimates chances of having affected child
• an provide couple with information on the
risk of genetic diseases

Genetic counsellor may discuss/advice on options:

• termination or therapeutic abortion
• therapies / treatments (e.g. gene therapy)
• financial implications of having affected child
• the effect of having affected child on existing siblings
• other ethical issues Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy
• To treat serious, common genetic diseases
• Caused by a single faulty, recessive allele
• By inserting normal, dominant allele
• Into affected cells of individual
• Via a vector (e.g. virus, liposome, or just naked DNA)
• Result = Host will have recombinant DNA

Aim:
Note: Gene editing in gametes or embryos is illegal
• To obtain functional normal polypeptide in humans! However, we can do it to somatic cells /
animal gametes and embryos to make GMOs
• Reduce symptoms of disorder
• Restore / enhance cellular functions
• Increase quality of life / survival

e.g. SCID, cystic fibrosis, Leber’s

congenital amaurosis (LCA), thalassaemia,
haemophilia
Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy
If faulty allele was dominant, gene therapy is very, very difficult because….
• It will still be expressed even when the normal, recessive allele is present
• We’ll need to knock out / remove / replace the faulty allele
→ Extremely difficult bcs it requires insertion of DNA into precise location in
genome (But we have a solution now! Refer to Extras – not in syllabus)

E.g. Huntington’s disease

• Caused by a faulty, autosomal dominant
allele of the huntingtin gene
• Dominant allele also affects tissues in
many parts of the body
→ Gene therapy only alters genotype
of a few targeted cells

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy
Procedure:
1. Obtain normal, dominant allele / cDNA from mRNA in cells of healthy person
2. Use gel electrophoresis + gene probe for identification
3. Use PCR to amplify DNA

4. Make recombinant DNA

• Use restriction enzyme to cut DNA and form complementary sticky ends
• Use DNA ligase to join cDNA with promoter / vector DNA
• Add human promoter upstream of target allele to ensure transcription in host

5. Insert normal allele into vector

• Virus vector / Liposome vector / naked DNA

6. Inject / spray into host

Updated on 12/8/21 by Beh SJ @behlogy

Types of Vectors
Gene Therapy
1. Virus vector
• Usually retroviruses
• Non-pathogenic
• Naturally inserts its viral DNA into
host genome
• Recognises specific cells

Problems:
• May cause side effects / allergies
• May be removed by immune system before it reaches target cells
• The virus may trigger an immune response which destroys the infected cells
• Most non-pathogenic viruses are not very good at getting into cells, so very
few cells receive the allele

• Short term effect as host cells have short lifespan

• Repeat treatments needed Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy
Problems:
• Retroviruses also cause random insertion of genes into host’s genome
• May insert its genes within another gene or within regulatory sequence
• May activate oncogenes / switch off tumour suppressor genes → cancer
• May have uncontrolled viral replication

Solution:
• Adeno-associated virus (AAV) does not
insert genes into host’s genome, not
passed on to daughter cells
• Lentiviruses can be modified to have
no uncontrolled viral replication
• Or use different vector!
Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy
Types of Vectors
2. Liposome vectors
• Small spheres of phospholipid
• Sprayed as an aerosol / Delivered using inhaler
• Liposome fuses with host cell surface membrane
• Trigger the immune response less than viruses

Problems:
• Not as effective in insertion as viruses
• Short term effect as host cells have short
lifespan
• Repeat treatments needed

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy
Types of Vectors
3. Naked DNA
• Very cheap
• Can be delivered via direct injection or gene gun
• No problems associated with using vectors
• Does not trigger immune response

Problems:
• Easily degraded
• Gene expression is very low

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: SCID
• Severe combined immunodeficiency (SCID) “Bubble boy”
https://www.youtube.com/watch?v=pJa6KVLwl9U

• Immune cells do not function properly

• Highly susceptible to infections
• B and T cells unable to make adenosine
deaminase (ADA)
• Due to faulty allele coding for the enzyme
• X-linked recessive allele

2. Introduce normal
1. Remove allele for ADA enzyme
T-lymphocytes using virus 3. Replace Problem:
T-lymphocytes Regular transfusions of
into body T cells are needed
(not a permanent cure)

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: SCID
Alternative, longer lasting method:
• Remove stem cells from bone
marrow
• Insert normal alleles into stem cells
using retroviruses
• Return stem cells into patient

Problem:
• Side effect = leukemia
• Due to random insertion of alleles
into cell’s genome

Solution:
• Use Adeno-associated viruses (AAV)

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: LCA
• Leber’s congenital amaurosis (LCA)
• Autosomal recessive eye disease
• Retina cells die off gradually
• Severe loss of vision at birth

• Delivery of dominant, normal allele using adeno-associated virus (AAV)

• Vectors are injected directly into the retina
• So retina cells can make functional protein and restore vision

Suitable for treatment using gene therapy bcs:

• Caused by recessive allele of a single gene
• Only need to get allele into a few cells
• Ease of access to affected area
• Only targets eye / no surgery needed
• Serious so worth the risk
Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy: Cystic Fibrosis
Chap 16 Recap
• Inherited genetic disease
• Faulty, autosomal recessive allele of the CFTR gene

Normal CFTR protein :

• Transmembrane protein at cell surface membrane
• Acts as chloride channel
• Has binding site for ATP
• Cl- moves out of cell via active transport
• Water is drawn out from cell
→ Normal / less viscous mucus formed
→ Easy removal by cilia

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: Cystic Fibrosis
Chap 16 Recap
Faulty CFTR allele:
a) Base deletion → faulty CFTR (most common)
b) Base substitution → STOP codon → incomplete CFTR

Faulty CFTR protein:

• No functional channels for Cl- ions
• Cl- ions do not move out
• Less water leaves cell
→ Formation of thick, sticky mucus
on cell surface membrane
→ Cannot be removed by cilia

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: Cystic Fibrosis
Chap 16 Recap
Symptoms:
• thick and sticky mucus produced at lungs
• mucus not moved effectively by cilia → mucus accumulates
• mucus traps bacteria → more infections
• reduced gaseous exchange → due to longer diffusion pathway
• difficulty in breathing, wheezing
• coughing → cause lungs to be scarred

• blocked pancreatic duct → reduced

digestion, damage of pancreatic |
tissues causes diabetes
• blocked sperm ducts / oviducts
→ reduced fertility

Updated on 12/8/21 by Beh SJ @behlogy

 Gene Therapy: Cystic Fibrosis
Traditional treatments
• Deals with symptoms only not causes
1. Thick mucus in lungs → physiotherapy, percussion therapy to loosen &
remove mucus easily
2. Recurrent bacterial infections → antibiotics
3. Reduced digestion → enzyme supplements

Gene therapy
• Insert normal allele for CFTR into lungs cells
→ Usually inhaled or sprayed
• Treats the cause rather than the symptoms
• No physiotherapy, antibiotics etc. needed
• Less time consuming than other treatments
• Effects are short-lived (few days)
and treatment needs repeating
• May have side-effects
Updated on 12/8/21 by Beh SJ @behlogy
 Gene Therapy: Cystic Fibrosis
Vectors commonly used:

1) Liposomes
• Sprayed into nose
• Not long lasting & only a few cells received
the normal allele

2) Adenoviruses
• Normal harmless
• Used to infect respiratory cells
• Not all cells take up virus
• Side effects due to viral infection

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Technology 4
MICROARRAYS
Function: Identify expressed genes by detecting mRNA
+ Distinguish btwn alleles of a gene
Updated on 12/8/21 by Beh SJ @behlogy
 Chapter Outline
Technology Function Application
4. Microarrays Identify expressed genes • Genetic screening
by detecting mRNA • Drug testing
OR • Identify genes that are
Distinguish btwn alleles switched on/off in diseases
of a gene

Updated on 12/8/21 by Beh SJ @behlogy

 Microarrays
• Function:

1) To distinguish btwn alleles of a gene

• Use DNA microarray
• Compares alleles found in genomes of 2 individuals/species

2) To identify expressed / transcribed genes

• Transcription of a gene produces mRNA
• So can assess gene expression by measuring mRNA levels

• Compares relative mRNA levels between 2 samples

• One acts as a control (normal cells)

Updated on 12/8/21 by Beh SJ @behlogy

 Microarrays
Components: A microarray chip

• Involves a “chip” with has >10 000 cells

• Gene probe is bound at known positions to a chip
• Gene probe = short lengths of single-stranded DNA
(ssDNA) complementary to allele/gene

• Many copies of one type of probe placed in

one cell of the microarray
• Diff cells has a diff type of gene probe
complementary to specific allele/gene
• So many genes can
be assessed at the same time!

Updated on 12/8/21 by Beh SJ @behlogy

 Microarrays
1A. To distinguish btwn alleles of a gene
Extract DNA from 2 samples
• Cut into fragments
• Denature DNA into ssDNA by heating it

OR

1B. To identify expressed / transcribed genes

Isolate all mRNA of cells of 2 samples
• Use mRNA as a template to form single-
stranded complementary DNA (cDNA)
• Use reverse transcriptase

Updated on 12/8/21 by Beh SJ @behlogy

 Microarrays
2. Label ssDNA using fluorescent ‘tags’
• One sample red, one sample green

3. Labelled ssDNA from both samples are

washed over microarray
• Allow ssDNA to hybridise / bind to the
complementary gene probes
• Unbound ssDNA will be washed off
• Bound DNA will not be washed off

Updated on 12/8/21 by Beh SJ @behlogy

Updated on 12/8/21 by Beh SJ @behlogy
Updated on 12/8/21 by Beh SJ @behlogy
 Microarrays
Applications:
• Genetic screening – can distinguish btwn alleles of a gene
• Drug testing – identify which genes the drugs have acted on
• Identify genes that are overexpressed/not expressed in diseases

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Technology 5 (Extra! Not in syllabus)
BIOINFORMATICS
Function: Collection, processing and analysis of biological info and
data using computer software
Updated on 12/8/21 by Beh SJ @behlogy
 Chapter Outline
Technology Function Application
5. Bioinformatics Collection, processing • Role in drug development
• DNA sequencing and analysis of biological E.g. against Plasmodium
(details not required) info and data using
computer software • Protein and DNA seq provide
evidence for evolution
Allows comparison of
DNA and protein
sequences

Updated on 12/8/21 by Beh SJ @behlogy

 Bioinformatics
• Interdisciplinary field
• Collection, processing and analysis of biological
information & data
• Using computer programs / software

• It is a store / database
→ Collection of huge-quantities of data (“big data”) from all over the world
→ From DNA sequencing, microarrays, other techniques
→ Can search online for DNA and amino acid sequences
→ And use that to model / predict tertiary structure of protein

• Function: To allow comparison of DNA and protein sequences

Updated on 12/8/21 by Beh SJ @behlogy

 DNA sequencing
• Determine the exact sequence of bases in genes

• Fully automated process

• Genomes of many species have been published
• Gives information about the location of genes
• Provides evidence for the evolutionary links
between organisms

E.g. Human Genome Project

• All 22+XY chromosomes have been sequenced
• Gives info about the location of genes

Updated on 12/8/21 by Beh SJ @behlogy

Bioinformatics

Updated on 12/8/21 by Beh SJ @behlogy

 Bioinformatics
Example of Databases available:
i) Ensembl – genomes of vertebrates
https://asia.ensembl.org/index.html

ii) UniProt – primary sequences & functions of proteins

https://www.uniprot.org/

iii) BLAST search tool – an algorithm to search the database

for target DNA or protein sequence
https://blast.ncbi.nlm.nih.gov/Blast.cgi

Updated on 12/8/21 by Beh SJ @behlogy

 Bioinformatics
Applications:
• Identify evolutionary relationships / genetic relatedness
• By comparing between genomes/protein seq of diff species
• Close similarities = recent common ancestry

• Compare between genes / proteins of humans and model organisms

• E.g. mouse, Drosophila fruit flies, yeast, C. elegans roundworm
• If target gene have similar sequence and protein have similar structure
• Model organism can be used to investigate when and where genes have
effect in humans

• Identify genome seq and proteins of pathogens

• E.g. Plasmodium, Staphylococcus areus
• Can help in vaccine/drug development

Updated on 12/8/21 by Beh SJ @behlogy

Genetic Technology 6 (Extra! Not in syllabus)
CRISPR-Cas9
Function: cut DNA at specific loci and allow genome editing
Updated on 12/8/21 by Beh SJ @behlogy
 Chapter Outline
Technology Function Application
6. CRIPSR-Cas9 Cut DNA at specific loci Cure viral diseases and genetic
(Extra! Not in syllabus) and allow genome diseases, even cancer!
editing

Updated on 12/8/21 by Beh SJ @behlogy

CRISPR-Cas9
• Inexpensive, much easier and very effective
• Bacterial system to protect itself against virus

Components:
• Cas9 = enzymes that cuts DNA
• gRNA = guide RNA that be complementary to
target DNA

Result: A double-stranded break is introduced

at specific loci
• Cell will try to repair the double-stranded
break but repair is error prone
• Extra bases are included → gene is disrupted
and deactivated

https://www.youtube.com/watch?v=4YKFw2KZA5o

Updated on 12/8/21 by Beh SJ @behlogy

CRISPR-Cas9
Potential:
• Technology is versatile and improving
→ Can combine with other enzymes in order to introduce specific mutations /
increase transcription / insert gene at specific location etc.
→ Can combine with GFP and act like a gene probe in situ

• Very cool experiments and research

→E.g. The bold plan to end malaria with a gene drive https://www.youtube.com/watch?v=P0HPHUzsHbI

• Cure viral diseases and genetic diseases, even cancer!

→ Can just inject CRISPR into body
→ Extend life expectancy

Potential Issues / Problems: https://www.youtube.com/watch?v=jAhjPd4uNFY

• Increased preselection → Designer babies
• Super soldiers and biological weapons
Updated on 12/8/21 by Beh SJ @behlogy
Chapter Outline
Technology Function Application
1. Polymerase Chain Clone and amplify DNA Too many…
Reaction (PCR)
• Role of Taq polymerase

Updated on 12/8/21 by Beh SJ @behlogy

 Chapter Outline
Technology Function Application
2. Recombinant DNA To make many copies of Large-scale production of human
technology / Gene Cloning a gene or protein proteins as drugs
E.g. insulin, factor VIII for haemophilia,
• Properties of plasmids adenosine deaminase for SCID
• Promoters
• Marker genes (e.g. Genetic engineering of
fluorescent genes / genes crop plants and livestock
coding for easily stained
substances) • Golden riceTM
• Role of restriction • GM salmon
endonucleases, reverse • Insect resistance
transcriptase and ligase E.g. Bt maize, Bt cotton
• Herbicide resistance
E.g. maize, cotton, tobacco, oil
seed rape

• Ethical and social implications

of using GMOs in food
production
Updated on 12/8/21 by Beh SJ @behlogy
 P5
Chapter Outline
Technology Function Application
3. Gel electrophoresis To separate fragments of Genetic fingerprinting /
• For protein and for DNA DNA according to length DNA profiling
• Probes OR • Paternity testing
To separate diff proteins • Criminal Investigations
according to
mass/charge Genetic screening
• Breast cancer (BRCA1, BRCA2)
Distinguish btwn alleles • Genes for haemophilia, SCA,
of a gene Huntington’s Disease and CF

Gene Therapy
• Vectors (viruses, liposomes,
naked DNA)
• SCID, inherited eye disease, CF
• Social and ethical
considerations

Updated on 12/8/21 by Beh SJ @behlogy

 Chapter Outline
Technology Function Application
4. Microarrays Identify expressed genes • Genetic screening
by detecting mRNA • Drug testing
OR • Identify genes that are
Distinguish btwn alleles switched on/off in diseases
of a gene

5. Bioinformatics and DNA Collection, processing • Role in drug development

sequencing and analysis of biological E.g. against Plasmodium
(Extra! Not in syllabus) info and data using
computer software • Protein and DNA seq provide
evidence for evolution
Allows comparison of
DNA and protein
sequences
6. CRIPSR-Cas9 Function: cut DNA at Cure viral diseases and genetic
(Extra! Not in syllabus) specific loci and allow diseases, even cancer!
genome editing

Updated on 12/8/21 by Beh SJ @behlogy