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1. What do you think are the objectives
of scientists in genetic engineering?

2. How are scientists able to realize

their objectives in genetic engineering?
 Genetic engineering is the direct
manipulation of an organism’s genes
using biotechnology. It covers different
kinds of technologies used to alter the
genomes that include the insertion of
genes from other individuals either the
same or from different species that
aims to produce or improve products.
 What Transgenic means
• 'Trans-' means “crossing from one place to another”

• The ‘genic’ means “genes”

• So it means that bits of genes from different living

things have been bolted together and spliced into
another organism to make a new one which does
something which the scientists want it to do.
Transgenic organisms
-also called genetically
modified organisms or
GMOs, are
organisms that are
created through the
modification of their
genomes (genetic
make-ups)
 Since ancient times the practice of
genetic engineering had begun. Artificial
selection is done to indirectly
manipulate genes focusing on the
physical traits among organisms.
Reference:
 Selective breeding is a process when
animals with desired characteristics are
mated to produce offspring with those
desired traits such as Angus cows are
bred to increase more meat.
 References: https://www.google.com/url?sa=i&url=https%3A%2F%2Fwww.broadheath.coventry.sch.uk%2Fscience-smsc-
morality%2F&psig=AOvVaw2NyqzP69yYJnB9qLlIbBF5&ust=1669019025358000&source=images&cd=vfe&ved=0CBAQjRxqFwoTCPiDq8SqvPsCFQAAAAAdAAAAABAD
 Hybridizations are when two
individuals with unlike characteristics are
crossed to produce the best in both
organisms. An example would be the
disease-resistant potato called the
Burbank potato.
 High resistance to diseases + High
yield production = Burbank Potato

Luther Burbank
Reference: https://visualdictionary.org/wp-content/uploads/2020/10/Hybrid-Animals-1.jpg
 Inbreeding
is a technique of breeding genetically
similar organisms to maintain desired
traits found in domestic and livestock.
Referrence: https://blogs.biomedcentral.com/on-biology/wp-content/uploads/sites/5/2015/09/inbreeding-in-dogs-
 Inbreeding Holstein
dairy cattle has led to
increased milk
production, but cattle
are more difficult to
breed.
 It involves
the artificial manipulation, modification, and
recombination of DNA or other nucleic
acid molecules to modify an organism or
population of organisms.
 Gene splicing
Gene splicing is the process of
chemically cutting DNA in order to add
bases to the DNA strand. The DNA is cut
using special chemicals called
restriction enzymes.
 Allergy-free milk
AgResearch, a company owned
by the New Zealand
government, engineered a cow
to produce milk without one of
the proteins many people are
allergic too, known as β-
lactoglobulin.
1. Isolation of Genes

Look for the

gene for a
specific trait that
you want from
the DNA donor.
 Plasmid
-a small, circular,
double-stranded
DNA molecule that
is distinct from a
Isolate Plasmid
cell's chromosomal
DNA.
2. Cutting

Donor DNA

Plasmid Sticky Ends

• Cutting cleavage of DNA by
Restriction Enzymes
• The restriction enzymes are
called molecular scissors
cutting the DNA at specific
target sequences
• These overhangs of the donor
DNA will be paired with the
other overhangs (vector DNA)
 2. Selection of an appropriate
vector/vehicle
• The most commonly used as vectors are
plasmids
• The plasmids are not part of the main cellular
genome but they carry genes that provide the
host cell with useful properties such as drug
resistance, mating ability and toxins
production
Restriction Enzymes
• DNA-cutting enzymes
DNA SPLICING
• process by which
the DNA of an
organism is cut and a
gene, perhaps from
another organism, is
inserted.
3. Ligation and Insertion
Ligation -(join together) of the gene of
interest (e.g. from animal) with the
vector. The resulting molecule is called
recombinant DNA ( composed of DNA
of 2 different sources)
 DNA Ligase

http://www.slic2.wsu.edu:82/hurlbert/micro101/pages/Chap10.html#Sticky_ended_cut

Ligation –rejoining cut fragments of DNA and

forming artificial recombinant molecules
 4. Transformation

Recombinant DNA introduced into bacterial cell

Bacterial
cell
Bacterial
chromosome

Recombinant DNA
 5. Expression
Bacterial cell reproduces by Binary Fisson
Bacterial cell produces the polypeptide
Coded for by the donor DNA
 Summary of Steps
Donor DNA
Plasmid

1. Cut with restriction

enzymes
Donor DNA
Sticky
Ends

2. Ligase bonds
sticky ends Recombinant DNA
together
 Gel electrophoresis
It is a laboratory method used to
separate mixtures of DNA, RNA, or
proteins according to molecular size. In
gel electrophoresis, the molecules to be
separated are pushed by an electrical
field through a gel that contains small
pores.
 • Recombinant DNA is made by
mixing DNA from two different
sources.

• Recombinant DNA technology use

to remove and insert genetic
sequences from and into other
sequences of another organism.
Reference: https://ars.els-cdn.com/content/image/3-s2.0-B9780128031094000076-f07-02-
9780128031094.jpg
an enzyme that cuts
DNA at particular
places
DNA molecule (often plasmid or
virus) that is used as a vehicle to carry
a particular DNA segment into a host
cell as part of a cloning or recombinant
DNA technique
 anorganism that harbours a
parasite and supplies it with
nutrients.

Reference: https://iastate.pressbooks.pub/app/uploads/sites/49/2021/04/Figure-1_Recombinant-DNA-is-made-from-combining-DNA-from-
different-sources-1024x522.png
 • Transformation Using a
PROCESSES Vector
USED IN
RECOMBINANT • Vectorless gene transfer
DNA
TECHNOLOGY
• Transduction
 1. Transformation using a vector

Vectors- are organisms that

Recombinant DNA may be
are normally harmless but
created through
may help spread the
transformation with the
infection by transferring
help of a vector, such as
the genetic material from
bacterial cells
one host to another
 1. Transformation using a vector
The plasmid contains a gene
A selected portion of the sequence that serves as
foreign DNA is inserted into bacterial origin of
a small, circular DNA replication. This is where
molecule called plasmids foreign DNA can be inserted
into the bacterial cell
It also contains a genetic
marker which makes it
possible to distinguish
bacteria that carry the
plasmid-containing foreign
DNA
• During transformation, a
restriction endonuclease
enzyme is used to cut the
piece of the donor DNA
• This enzyme cleaves the
DNA at the phosphate sugar
bond and thus sticky ends
are created
• Sticky ends are areas in the
DNA where bases are ready
to be paired
• Then an enzyme known as DNA ligase is used
to insert the donor DNA into the vector. It seals
the sticky ends by joining the phosphate and
sugar bonds in the DNA
• The inserted DNA contains genetic marker for
identification
• The recombinant DNA is then inserted into the
bacterial cell, such as E.coli
• After transformation, the culture is treated
with an antibiotic.
• Those that have been transformed will be the
only ones to survive because they carry the
resistance gene
 Electroporation- temporary
holes are formed in the
2. plasma membrane of the host
Vectorless cell by applying a significant
gene amount of electricity in the
culture medium. This enables
transfer the entry of the foreign DNA
via the pores
 Protoplast fusion-cells are
2. treated with chemicals to
initiate recombination
Vectorless
gene • In this process, bacterial cell
transfer walls are digested, turning
cells into protoplasts
• These protoplasts are treated with
polyethylene glycol to allow them to fuse,
creating a random recombination of genes
• The resulting recombinant cell will now grow a
new cell wall
 Microinjection

• The host cell is immobilized

by applying mild suction
with a blunt pipette. The
foreign gene is then
injected with a
microinjection needle, thus
creating recombinant DNA
 • The host cell is bombarded with
Particle Gun for tungsten particles coated with the
Recombination foreign DNA.
• This process is used in the field of
agriculture
• The farmers use this method to
genetically modify plants to make
them highly resistant to insects and
other pests and develop crops that
cam survive extreme weather
conditions
 • It is the process wherein
genetically engineered
bacteriophages-viruses that
Transduction parasitize bacteria are
introduced into the cell to
create the desired
recombinant DNA
 Applications of recombinant DNA in
genetic engineering are: For the
production of vaccines like the hepatitis B
vaccine. Production of transgenic plants
with improved qualities like insect and
drought resistance and nutritional
enrichment. Therapeutic protein
production like insulin.
INTRODUCTION

 In order to survive, man has successfully

domesticated selected plants and
animals. He has taken an active part in
choosing desired traits of plants and
animals.
INTRODUCTION

 Traits that were considered valuable were sought out and propagated.
 The process involved may include classical breeding practices such as
controlled pollination of plants, and the mating of animals with desired
traits.
In today’s modern science, molecular biology techniques are being
employed in the insertion and expression of proteins in different organisms
for various purposes.
 KOBE / WAGYU BEEF
BEEF WITH GOOD FAT
DISTRIBUTION
 GUAPPLE
LARGE SIZED GUAVA
HUMAN INSULIN-PRODUCING
BACTERIA
 FLAVR-SAVR
Delayed-ripening Tomatoes
MACAPUNO TRAIT IN COCONUT
CLASSICAL BREEDING

 Classical breeding practices

focus on the mating of
organisms with desirable
qualities.
GENETIC ENGINEERING

 It involves the use of molecular techniques to modify the traits of

a target organisms.The modification of traits involve:
a. Introduction of new traits into an organism
b. Enhancement of a present trait by increasing the expression of
the desired gene
c. Enhancement of a present trait by disrupting the inhibition of
the desired gene’s expression.
GENERAL OUTLINE
RECOMBINANT DNA
 Ways in which these plasmids may be introduced into host organism.
WAYS
BIOLISTICS: a gene gun is used to fire DNA-coated pellets on
plant tissues. Cells that survive bombardment, and are able to take up
the expression plasmid coated pellets and acquire the ability to
express the designed protein.
 Ways in which these plasmids may be introduced into host organism.
WAYS
PLASMID INSERTION BY HEAT SHOCK TREATMENT
• It is a process used to transfer plasmid DNA into bacteria.
• The target cells are pre-treated before the procedure to increase
the pore sizes of their plasma membranes.
• This pretreatment (usually with CaCl2) is said to make the cells
“competent” for accepting the plasmid DNA.
 Ways in which these plasmids may be introduced into host organism.
WAYS
PLASMID INSERTION BY HEAT SHOCK TREATMENT

• After the cells are made competent, they are incubated with the
desired plasmid at about 4°C for about 30min.

• The plasmids concentrate near the cells during this time.

Afterwards, a “Heat Shock” is done on the plasmid-cell solution
by incubating it at 42°C for 1 minute then back to 4°C for 2
minutes.
 Ways in which these plasmids may be introduced into host organism.
WAYS
PLASMID INSERTION BY HEAT SHOCK TREATMENT

• The rapid rise and drop of temperature is believed to increase

and decrease the pore sizes in the membrane.

• The plasmid DNA near the membrane surface are taken into the
cells by this process. The cells that took up the plasmids acquire
new traits and are said to be “transformed”.
 Ways in which these plasmids may be introduced into host organism.
WAYS
ELECTROPORATION
• This technique follows a similar methodology as Heat Shock
Treatment, but the expansion of the membrane pores is done
through an electric “shock”.
• This method is commonly used for insertion of genes into
mammalian cells.
GENETICALLY MODIFIED ORGANISMS (GMO)

 With the ability to insert gene sequences, comes the possibility of

providing new traits for these target organisms.
 This has allowed the development of GMOs. Some of these genetic
modifications promise higher product yield for their targets. These
include the Flavr-Savr Tomato and Bt-Corn.
GENETICALLY MODIFIED ORGANISMS (GMO)

 The Flavr-Savr (“Flavor Savor”) tomato was the first genetically

modified organism that was licensed for human consumption. The trait
modified in this tomato is its ripening process.
 A gene for an enzyme that causes the degradation of pectin in the cell
walls (i.e. polygalacturonase) normally softens the fruit as it ripens.
 In Flavr Savr tomatoes, an inhibitor (i.e. antisense RNA) disrupts the
expression of this gene, thereby delaying the softening of the fruit and
extending the time it may be kept in storage and transported to markets.
GENETICALLY MODIFIED ORGANISMS (GMO)

 Bt-Corn was developed to incorporate the production of a toxin (i.e. Bt-

endotoxin) from Bacillus thuringiensis in corn plants.
 This toxin results in the death of pests that feed on these plants like the
corn borer larvae. The toxin has been shown to be selective for
Lepidoptera larvae and is non-toxic to humans, mammals, fish and birds.
GENETICALLY MODIFIED ORGANISMS (GMO)

 The selective toxicity of the toxin allows its use in food crops.
 The introduction of the toxin is believed to increase crop production due
to decreased losses from pest infestation. The same technology has been
applied in the Philippines for the development of Bt-Eggplant
GENETICALLY MODIFIED ORGANISMS (GMO)

 Despite the proposed benefits of GMOs, some people have raised their
concerns regarding the consumption of these modified foods. While most
of the products are tested for safety, concerns are raised for the
possibility of not being able to detect hazards that are present, but are
currently undetectable by today’s current technology.
GENETICALLY MODIFIED ORGANISMS (GMO)

 Because of these issues, manufacturers are urged to provide labels that

notify consumers of GMO presence in their products. While GMOs are
believed to be safe when licensed by the food regulatory agencies, it is
believed that the consumers must be provided with enough information
to make their own choices regarding their use.
1. PRODUCTION OF TRANSGENIC PLANTS

 By utilizing the tools and techniques of genetic engineering, it is

possible to produce transgenic plants or genetically modified
plants. Many transgenic plants have been developed with better
qualities like resistance to herbicides, insects, or viruses or with
the expression of male sterility, etc
2. PRODUCTION OF TRANSGENIC ANIMALS
 By the use of rec DNA technology, desired genes can be inserted into
the animal so as to produce the transgenic animal.
 The method of rec DNA technology aids the animal breeders to increase
the speed and range of selective breeding in the case of animals.
 It helps for the production of better farm animals to ensure more
commercial benefits. Another commercially important use of transgenic
animals is the production of specific proteins and pharmaceutical
compounds. Transgenic animals also contribute to studying the gene
functions in different animal species. Biotechnologists have successfully
produced transgenic pigs, sheep, rats, and cattl
3. PRODUCTION OF HORMONES

 Production of Hormones By the advent of techniques of

rec DNA technology, bacterial cells like E.coli are utilized
for the production of different fine chemicals like insulin,
somatostatin, somatotropin, and endorphin. Human
Insulin Hormone, I
 .e., Humulin, is the first therapeutic product that was
produced by the application of rec DNA technology.
4. PRODUCTION OF VACCINES
 Vaccines are the chemical preparations containing a pathogen in an attenuated (or
weakened) or inactive state that may be given to human beings or animals to confer
immunity to infection.
 A number of vaccines have been synthesized biologically through recDNA technology;
these vaccines are effective against numerous serious diseases caused by bacteria,
viruses, or protozoa.
 These include vaccines for polio, malaria, cholera, hepatitis, rabies, smallpox, etc.
 The generation of DNA vaccines has revolutionized the approach to the treatment
of infectious diseases.
 DNA-vaccine is the preparation that contains a gene encoding an immunogenic
protein from the concerned pathogen
5. BIOSYNTHESIS OF INTERFERON

 Interferons are the glycoproteins that are produced in very minute amounts by
the virus-infected cells.
 Interferons have antiviral and even anti-cancerous properties.
By the recDNA technology method, the gene of human fibroblasts (which
produce interferon's in human beings) is inserted into the bacterial plasmid.
 These genetically engineered bacteria are cloned and cultured so that the gene
is expressed and the interferons are produced in relatively high quantities. This
interferon, so produced, is then extracted and purified.
6. PRODUCTION OF ANTIBIOTICS

 Antibiotics produced by microorganisms are very effective against

different viral, bacterial, or protozoan diseases.
 Some important antibiotics are tetracycline, penicillin, streptomycin,
novobiocin, bacitracin, etc.
 The recDNA technology helps in increasing the production of antibiotics
by improving the microbial strains through modification of genetic
characteristics.
7. PRODUCTION OF COMMERCIALLY IMPORTANT CHEMICALS

 Various commercially important chemicals can be produced

more efficiently by utilizing the methods of rec DNA
technology. A few of them are the alcohols and alcoholic
beverages obtained through fermentation, organic acids like
citric acid, acetic acid, etc., and vitamins produced by
microorganisms
8. APPLICATION IN ENZYME ENGINEERING

 As we know that the enzymes are encoded by genes, so if there

are changes in a gene, then definitely the enzyme structure also
changes. Enzyme engineering utilizes the same fact and can be
explained as the modification of an enzyme structure by
inducing alterations in the genes which encode for that
particular enzyme.
9. PREVENTION AND DIAGNOSIS OF DISEASES

 Genetic engineering methods and techniques have greatly

solved the problem of conventional methods for the diagnosis
of diseases. It also provides methods for the prevention of a
number of diseases like AIDS, cholera, etc. Monoclonal
antibodies are useful tools for disease diagnosis.
 Monoclonal antibodies are produced by using the technique
called hybridoma technology.
10. GENE THERAPY

 Gene therapy is undoubtedly the most beneficial area of genetic

engineering for human beings.
 It involves the delivery of specific genes into the human body to
correct the diseases. Thus, it is the treatment of diseases by
transfer and expression of a gene into the patients' cells so as to
ensure the restoration of a normal cellular activity.