11

GENERAL
BIOLOGY II

3 Quarter
rd

RECOMBINANT DNA
TECHNOLOGY
(WEEK 1)
GENERAL BIOLOGY II
Content Standards:
The learners demonstrate an understanding of Recombinant DNA

Performance Standards:
The learners should be able to make a research paper/ case study / Poster on genetic
diseases.

Most Essential Learning Competencies:

1. Outline the processes involved in genetic engineering (STEM_BIO11/12-IIIa-b-6)

2. Discuss the application of recombinant DNA (STEM_BIO11/12-IIIa-b-7)

Lesson OUTLINE THE PROCESSES

INVOLVED IN GENETIC
1 ENGINEERING

Genetic engineering is a process of making changes on the genetic code of an organism.

Its goal is to add one or more new traits that are not normally found in that organism. Through
advance studies in the structure of DNA and its chemical properties, scientists have been able to
employ different techniques to extract, cut, and make unlimited copies of DNA.
DNA recombination is a process of modifying the genes of organisms for practical
purposes. It is done when a piece of DNA is combined with another DNA from another source. The
resulting genetic product is called recombinant DNA. With this process, organisms get to have
traits that are not normally found in their species.

Brief History of Recombinant DNA Technology

In the late 1960’s, Stewart Linn and Werner Arber discovered restriction enzymes in E. coli,
which are known as endonucleases. Endonucleases cut DNA at specific sites where there are
adjacent base sequences. They also create sticky ends on the cut DNA sites, allowing certain DNA
fragments to be joined together. In 1969, Herbert Boyer isolated restriction enzyme EcoRI from E.
coli. That cleaves the DNA between G and A in the base sequence GAATT. In 1970,
Howard Temin and Davin Baltimore independently discovered the enzyme reverse transcriptase
from retroviruses. This enzyme was used to construct a
DNA called complementary DNA (cDNA) from any mRNA. In, 1972 David Jackson, Robert Symons
and Paul Berg successfully generated rDNA molecules. They allowed the sticky ends of
complementary DNA by using an enzyme DNA ligase. In 1973 for the first time S.Cohen and
H.Boyer developed a recombinant plasmid (pSC101) which after using as vector replicated well
within a bacterial host. In, 1975, Edwin Southern developed a method for detection of specific DNA
fragments for isolation of a gene from complex mixture of DNA. This method is known as the
Southern blotting technique. In 1976, first prenatal diagnosis by using gene specific probe. In
1977, methods for rapid DNA sequencing, discovery of split genes and somatostanin by rDNA. In
1979, Insulin synthesized by using rDNA; first human viral antigen. In 1981, foot and mouth
disease viral antigen cloned. 1981 – Foot and mouth disease viral antigen cloned. 1982 –
Commercial production of coli of genetically engineered human insulin, Isolation, cloning and
characterization of human cancer gene. 1983 – Engineered Ti-plasmid used to transform plants.
1985 – Insertion of cloned gene from Salmonella into tobacco plant to make resistant to herbicide
glyphosphate; Development of PCR technique. 1986 – Development of gene gun.1989 – First field
test of genetically engineered virus (baculovirus) that kills cabbage looper caterpillars. 1990 –
Production of first transformed corn. 1991 - Production of first transgenic pigs & goats,
manufacture of human haemoglobin and 1st test of gene therapy on human cancer patients.
In1994, first genetically engineered whole food approved for sale and human monoclonal
antibodies produced in genetically engineered mice.
Technologies and Tools Used in Recombinant DNA Technology
Molecular biologists have developed different technologies and tools that allow them to
study and manipulate DNA molecules. The processes that will be discussed here are artificial
selection, cloning, gene splicing, gel electrophoresis, DNA sequencing and polymerase
chain reaction (PCR).

ARTIFICIAL SELECTION - breeders choose which organism to mate to produce offspring

with desired traits. They cannot control what genes are passed. When they get offspring with the
desired traits, the maintain them.
Three types of artificial selection:
A Selective breeding: when animals with desired characteristics are mated to produce
offspring with those desired traits. Passing of important genes to next generation.
Example:
• Champion race horses, cows with tender meat, large juicy oranges on a tree.
• For example people breed dogs for specific purposes.
• Dachshund were once bred to hunt badgers and other burrowing animals.
• They must be small to fit into the animals hole in the ground.

Selective breeding occurs when you choose the best male and female to breed. This allows you
to fine tune and control the traits. The offspring or babies will then have the best traits. Then you
continue to breed those organism with the best traits, those traits will be maintained.

B. Hybridizations: two individuals with unlike characteristics are crossed to produce the best
in both organisms.
Example:
• Luther Burbank created a disease resistant potato called the Burbank potato. He
crossed a disease resistant plant with one that had a large food producing capacity.
Result: disease resistant plant that makes a lot of potatoes. Hybridization
• Liger: Lion and Tiger Mix
• Grapple: Grape plus apple mix. The fruit tastes like grapes and looks like apple.

C. Inbreeding: breeding of organism that genetically similar to maintain desired traits.

Example:
• Dogs breeds are kept pure this way. It’s how a Doberman remains a Doberman. It keeps
each breed unique from others.
Risk: since both have the same genes, the chance that a baby will get a recessive genetic
disorder is high. Risks: blindness, joint deformities.

CLONING: creating an organism that is an exact genetic copy of another.

There are human clones in our school. Identical twins are naturally created clones. Clone:
group of cells or organisms that are genetically identical because of asexual reproduction.
They will have the same exact DNA as the parent.

How is cloning done?

• A single cell is removed from a parent organism.
• An entire individual is grown from that cell.
• Remember one cell has all the DNA needed to make an entire organism.
• Each cell in the body has the same DNA, but cells vary because different genes are turned
on in each cell.

Example:
Dolly the Sheep
• Dolly was the first mammal cloned. She had the same exact DNA as her mother and had
no father. Cloning is a form of asexual reproduction. Only one genetic parent.
 The Process of Cloning
Benefits of cloning:
1. You can make exact copies of organisms with strong traits.
2. Increase food supply
3. Medical purposes: clone organs for transplants.
4. Bring back or stop species from going extinct.
Risks of cloning:
1. Decreases genetic diversity
2. If one of your clones gets a disease, they all get it: same immune system.
3. Inefficient: high failure rate: 90%+
4. Expensive

GENE SPLICING: DNA is cut out of one organism and put into another organism
• A trait will be transferred from one organism to another.
• For example: the human insulin gene can be removed from a human cell.
• It can be put into a bacterial cell.
• The bacterial will now make human insulin.

Benefits:
• insulin is cheaper
• There are no side effects because it is
human insulin.
• We once used pig insulin but there are
side effects and it more expensive.

How are genes cut for gene splicing?

• A bacterial plasmid is used.
• Plasmid: circular DNA in a bacteria
cell.
• It is very simple and easy to
manipulate.

• A restriction enzyme: enzyme that cuts the DNA at a specific code.

• There are thousands of restriction enzymes.
• Each cuts DNA at a different sequence.
• Some look for GGCC and cut in between the G and C.
• Every time GGCC is found in the DNA it is cut by the restriction enzyme
DNA Code:
• TTATGGCCATACGGCCTT
• AATACCGGTATGCCGGAA

• TTATGGCCATACGGCCTT
• AATACCGGTATGCCGGAA

• TTATGG CCATACGG CCTT

• AATACC GGTATGCC GGAA

• This DNA segment was cut twice creating three fragments.

• Since every one is different, we all have a different amount of times GGCC is found. DNA
may be cut seven times.

How is gene splicing done?

1. A restriction enzyme cuts the insulin gene out of the human DNA.
2. A plasmid is removed from a bacteria and cut with a restriction enzyme

3. The human gene is place into the bacteria plasmid

4. The plasmid is placed back into the bacteria.
• The cell now has directions (DNA) to make insulin.
• That's exactly what it does.
• Its human insulin, bacteria do not make insulin on their own.

Plasmid with
insulin gene

This is called transformation: when a gene from one organism is transferred to different
organism. The organisms that have DNA transferred to them are called transgenic organisms.
• trans: means different,
• genic: refers to genes
• Genetic engineering has given rise to a new technological field called biotechnology
(technology of life).

GEL ELECTROPHORESIS: a
technique used to compare DNA from two or
more organisms.
Why compare DNA:
1. Find your baby’s daddy
2. Who committed a crime.
3. How closely species are related.

How is electrophoresis done?

A. The DNA is cut into fragments with a restriction enzyme.
B. The cut DNA is then put into the wells of a machine filled with gel.
• The gel is spongy and the DNA squeezes through
the pores.
C. The machine is plugged in and the fragments get
separated based on their size.
• The smaller fragments move further than the
large.
• Electricity provides the energy
• Why does DNA move?
• DNA has a negative charge.
• When the machine is plugged it, its moves
towards the positive pole created by the
electricity

DNA SEQUENCING: this is a method used

to provide the identify and order of
nucleotides in a DNA strand. Small, single
stranded pieces of DNA are placed in test
tubes with an enzyme that can make a
complementary DNA strand by using the
original DNA strand as a template. A supply
of the four nucleotide bases found in DNA
is then added, along with a small amount of
one of the bases that has been labeled with
fluorescent dyes.
Polymerase Chain Reaction (PCR): Information must be acquired for either its detection or
expression in a given organism, once a desired trait is chosen. This is important in detecting
diseases or infectious agents. The goal of PCR is to amplify specific DNA sequences. To make copies
of DNA, DNA is heated to separate its strands and then it is cooled down to allow the primers to
bind to the single- strand DNA.
The primers are short type of DNA strands that provide a place for the DNA polymerase to
start working. As the polymerase starts working, new strands of the separated DNA are formed.
Continuous heating and cooling allow further separation of DNA and formation of new DNA
strands, respectively, creating millions of copies of the DNA segments. Figure below shows how
PCR can create copies of DNA strands.

PCR Applications
PCR might be utilized to recognize the presence of an ideal gene/quality in a living being.
Contingent upon the preliminary plan, the normal item may speak to just a particular district of
the quality or the whole quality itself. The main case is valuable for location of the quality, or the
recognition of living beings with that particular quality inside a sample. The subsequent case is
helpful for the enhancement of the whole quality for inevitable articulation in different living
beings. The immediate enhancement/duplicating of a full quality is important for the cycle for
"cloning" that gene.

Lesson APPLICATIONS OF RECOMBINANT

2 DNA TECHNOLOGY

Application of Recombinant DNA

1. Production of Transgenic Plants:
By using the apparatuses and strategies of hereditary designing it is conceivable to deliver
transgenic plants or the hereditarily altered plants. Numerous transgenic plants have been
created with better characteristics like protection from herbicides, creepy crawlies or infections
or with articulation of male sterility and so forth.

2. Production of Transgenic Animals:

By the utilization of rec DNA technology, wanted qualities can be embedded into the
creature to deliver the transgenic creature. The technique for rec DNA technology helps the
creature raisers to speed up and scope of specific reproducing if there should arise an
occurrence of creatures. It helps for the creation of better livestock to guarantee more business
benefits. Another economically significant utilization of transgenic creatures is the creation of
specific proteins and drug mixes. Transgenic creatures likewise contribute for considering the
quality capacities in various creature species. Biotechnologists have effectively delivered
transgenic pigs, sheep, rodents and steers.

3. Production of Hormones
Bacterial cells like E.coli are used for the creation of various fine synthetic substances like
insulin, somatostatin, somatotropin and pendorphin by the appearance of strategies of rec DNA
technology,. Human Insulin Hormone i.e., Humulin is the principal helpful item which was
delivered by the use of rec DNA technology.

4. Production of Vaccines
Vaccines are the synthetic arrangements containing a microbe in constricted (or debilitated)
or idle express that might be given to people or animals to give invulnerability to contamination.
Various immunizations have been incorporated organically through rec DNA technology; these
antibodies are powerful against various genuine sicknesses brought about by microbes,
infections or protozoa. These incorporate immunizations for polio, jungle fever, cholera,
hepatitis, rabies, smallpox, and so forth. The age of DNA antibodies has altered the
methodology of treatment of irresistible sicknesses. DNA-immunization is the planning that
contains a quality encoding an immunogenic protein from the concerned microbe.

5. Biosynthesis of Interferon
Interferon’s are the glycoproteins which are created in exact moment sums by the infection
contaminated cells. Interferon's have antiviral and even enemy of dangerous properties. By
recombinant DNA technology strategy, the quality of human fibroblasts (which produce
interferon's in individuals) is embedded into the bacterial plasmid. These hereditarily designed
microbes are cloned and refined with the goal that the quality is communicated and the
interferons are created in genuinely high amounts. This interferon, so delivered, is then
separated and filtered

6. Production of Antibiotics
Antibiotics delivered by microorganisms are exceptionally powerful against various viral,
bacterial or protozoan ailments. Some significant anti-toxins are tetracyclin, penicillin,
streptomycin, novobiocin, bacitracin, etc. recombinant DNA technology helps in expanding the
creation of anti-infection agents by improving the microbial strains through change of
hereditary attributes.

7. Production of Commercially Important Chemicals

Various industrially significant synthetic substances can be delivered all the more
proficiently by using the techniques for rec DNA technology. A couple of them are the alcohols
and mixed drinks got through aging; natural acids like citrus extract, acidic corrosive, and so
on and nutrients delivered by microorganisms.

8. Application in Enzyme Engineering

As we realize that the chemicals are encoded by genes, so in the event that there are changes
in a quality, at that point unquestionably the catalyst structure likewise changes. Catalyst
building uses a similar truth and can be clarified as the adjustment of a chemical structure by
inciting changes in the qualities which encode for that specific protein.

9. Prevention and Diagnosis of Diseases

Genetic building strategies and procedures have enormously tackled the issue of regular
techniques for determination of illnesses. It additionally gives strategies to the. counteraction
of various sicknesses like AIDS, cholera, and so on. Monoclonal antibodies are helpful
instruments for illness finding. Monoclonal antibodies are delivered by utilizing the method
called hybridoma innovation.

10. Gene Therapy

Gene treatment is without a doubt the most helpful zone of hereditary building for
individuals. It includes conveyance of explicit qualities into human body to address the
 sicknesses. Hence, it is the treatment of illnesses by move and articulation of a quality into the
patients' cells to guarantee the rebuilding of a typical cell action.

11. Practical Applications of Genetic Engineering

Recombinant DNA technology has an enormous degree in Research and Experimental
examinations It is applied for:
• Localizing explicit genes.
• Sequencing of DNA or genes.
• Study of component of gene guideline.
• Molecular examination of different infections.
• Study of changes in DNA, and so on.

12. Applications in forensic science

The uses of rec DNA technology (or hereditary designing) in legal sciences generally rely
upon the method called DNA profiling or DNA fingerprinting. It empowers us to distinguish any
individual by dissecting his hair roots Wood stains, serum, and so forth. DNA fingerprinting
additionally assists with tackling the issues of parentage and to recognize the lawbreakers.

13. Biofuel Production

Biofuels are gotten from biomass and these are sustainable and savvy. Hereditary designing
assumes a basically significant part in an advantageous and enormous scope creation of
biofuels like biogas, bio-hydrogen biodiesel bio-ethanol., and so forth. Hereditary building
assists with improving creatures for getting higher item yields and item resistance.

14. Genetically steady high creating microorganisms are being created by utilizing present day
rDNA methods, which help in a proficient creation of bioenergy.

15. The vitality crop plants are those plants which utilize sunlight-based vitality in a superior
manner for creation of biomass. Hereditary enhancements of these vitality crop plants
enormously help for snappy and high Product on of biomass which thusly lessens the
biofuel creation cost. The aging microorganisms which are used for biogas creation are
improved at the hereditary level for accomplishing better outcome.

16. Environment Protection

Genetic engineering makes its commitments to nature security in different manners.
Generally critical to make reference to are the new methodologies used for squander
medicines and bioremediation Environment assurance implies the protection of assets and
thus to restrict the debasement of condition.

WHAT I HAVE LEARNED

Test I. Match each term in column A with its definition in column B. Write the letter of
the correct answer.
COLUMN A COLUMN B
_____1. Plasmids a. Cuts the DNA into fragments
_____2. Sticky Ends b. Circular DNA molecule of bacteria
_____3. DNA Ligase c. Used to insert DNA of interest to vector
_____4. Transformation d. Area of DNA where bases are ready to be paired
_____5. Restriction e. Recombinant DNA technology with the help of a
Enzyme vector gene.
Test II. In your own words, define or describe the functions of each term used in
recombinant DNA technology. (5 pts. Each)

1. Recombinant DNA
________________________________________________________________________________________
________________________________________________________________________________________
 2. DNA Ligase
________________________________________________________________________________________
________________________________________________________________________________________
3. Restriction Enzymes
________________________________________________________________________________________
________________________________________________________________________________________
4. PCR
________________________________________________________________________________________
________________________________________________________________________________________
5. Sticky Ends
________________________________________________________________________________________
________________________________________________________________________________________

Test III. Encircle the letter of the correct answer.

1. Which gene transfer technique involves the direct injection of DNA into a cell lacking that
DNA sequence?
a. Gel electrophoresis c. Electroporation
b. Microinjection d. Particle Gun

2. At what level are gene products manipulated in genetic engineering?

a. Protein b. Amino acid c. RNA d. DNA

3. Which of the following best defines the function of Agrobacterium tumefaciens?

a. It can cause blindness in humans.
b. It can be used to introduce DNA to plants.
c. It can cause weight loss in humans.
d. It is used for recombinant DNA in fungi.

4. What are vectors?

a. Molecules that help in replication
b. Molecules that carry foreign DNA into cells
c. Molecules that degrade nucleic acids
d. Molecules that protect host cells from foreign DNA

5. One of the application of Recombinant DNA that contain genes from other organisms, that
are now important part in the field of agriculture is called __________.
a. Transgenic plants c. Transduction
b. Insulin d. Gene Splicing

Test IV. Answer the following questions briefly. (5 pts each)

1. What is the significance of recombinant DNA technology? Cite specific examples from
different fields.
________________________________________________________________________________________
________________________________________________________________________________________
2. Describe the advantages and disadvantages of recombinant DNA technology.
________________________________________________________________________________________
________________________________________________________________________________________
3. Identify the risks and benefits of consuming food that came from GMO’s. Do you think
there is a need for stricter control in their production? Why or why not?
________________________________________________________________________________________
________________________________________________________________________________________
PERFORMANCE TASKS NO. 1 (Week 1)
Make a research paper/case study/poster on genetic diseases. You can make use of any
available paper for your poster.

RUBRIC FOR POSTER MAKING:

REFERENCES

Javier, M.A., (2017). DIWA Senior High School Series: General Biology 2. DIWA Learning
Systems Inc., pp 96 - 112

S. A. Shinde*, S. A. Chavhan, S. B.Sapkal, V. N. Shrikhande, (2018).Recombinant DNA

Technology and its Applications: A Review. International Journal of MediPharm Research,
Vol.04, No.02, pp 79-88. Retrieved from
https://www.researchgate.net/publication/327111668_Recombinant_DNA_Technology_an
d_its_Applications_A_Review on 14, September 2020.

Teaching Guide for Senior High School-General Biology 2 (2016) Commission on Higher Education,
pp. 47-59

Prepared:

TERESA ELIEZEL B. COLLONG

Teacher III

Checked:

Rosita A. Elopre
Master Teacher I

Noted:

Ma. Victoria C. Vivo Ed.D

Principal IV