RECOMBINANT DNA - Combination of DNA from different species.

- With this technology, a gene or multiple genes can be identified, cut, and inserted into the
genome of an organism.
- THREE MAIN TOOLs
1. ENZYMES (restriction) (Synthesize) (Polymerases) and (ligases) DNA –
- Restriction enzymes play an important role in this technology. The restriction enzyme will cut at
a specific site within the DNC molecule called a restriction site. Usually, the restricted enzymes
will produce sticky ends in the DNA sequence that will help in binding specifically to the desired
gene.
2. VECTORS – they will carry the desired gene. They are considered as the final vehicles that carry
genes of interest into the host organism. The most used vectors are plasmids and
bacteriophages.
3. HOST ORGANISM- is the cell in which the recombinant is introduced. To date, host organisms
include bacteria, fungi, and animal cells.
- To introduce vectors into the host, techniques involving microinjection, biolistic, gene gun,
alternate cooling and heating and calcium phosphate ions have been used.

FIVE STEPS IN RECOMBINANT DN

1. Cutting the desired DNA by

restriction sites.
2. Amplifying the gene copies PCR.
3. Insert genes into vectors.
4. Transferring the vectors into the host
organism
5. Obtaining the product of
recombinant genes.

CHECK YOUR UNDERSTANDING

1. They are considered as the final vehicles that carry genes of interest into the host organism.
2. It helps cut (restriction enzymes), synthesize (polymerases), and bind (ligases) DNA.
3. It is the cell in which the recombinant DNA is introduced.
4. This enzyme cuts the desired DNA and the vector.
5. What are the five steps in recombinant DNA?

GENETIC ENGINEERING

Genetic – Study of heredity

Genes – the carrier of organism traits

Ribonucleic Acid – It is a unique polymer that can accelerate a chemical reaction and bind specific
proteins or small molecules.

Deoxyribonucleic Acid –is called the blueprint of life. It contains the genetic code of an organism.

Mutation – it is any change of base–pair sequence of genetic materials.

Polymerase – an enzyme that assembles DNA and RNA molecules and synthesizes long chains of
polymers or nucleic acids.

A genetically Modified Organism- Is an organism or microorganism whose genetic materials have been
altered to contain a segment of DNA from another organism.

Genetic Engineering- is a modern biotechnology that produces transgenic or genetically modified crops
of organisms. It is a method that moves genes from one specie into another.

Genes are made up of a sequence of RNA and DNA. They carry an organism’s trait and are fixed in one
position inside a chromosome.

Polymerase Chain Reaction- Duplication of DNA. Makes possible to the detention of pathogens.
Pathogens – Can cause disease (virus) bacteria.

Thermal cycles – carries out temperature cycle into a test tube.

The DNA polymerase typically used in PCR is called Taq polymerase, after the heat-tolerant bacterium
from which it was isolated (Thermus aquaticus)

The Steps of PCR

1. Denaturation (96 Celsius): Heat the reaction strongly to separate, or denature, the DNA strands.
This provides a single-stranded template for the next step.
2. Annealing (55-65 Celsius): Cool the reaction so the primers can bind to their complementary
sequences on the single-stranded template DNA.
3. Extension (72 Celsius): Raise the reaction temperatures so Taq polymerase extends the primers,
synthesizing new strands of DNA.

Molecular Scissors/ Restriction Enzyme

Insulin- is made by bacteria that have the human gene for insulin.

The easiest way to isolate a gene is to cut it out and clone the gene into small piece of bacterial DNA
called plasmid.

How does gene manipulation take place -?

1. DNA of the organism whose gene is to be altered is isolated.

2. After isolating the DNA, a segment that corresponds to the quality we want to change is cut.

Gel Electrophoresis – is a technique used to separate DNA fragments (or other macro molecules, such as
RNA and protein) based on their size. It involves running a current through a gel containing the
molecules of interest.

DNA Fingerprinting – After the DNA has been digested with one or more restriction enzymes, it is
separated by size using gel electrophoresis. By comparing the size of DNA fragments produced by
restriction digest, the DNA fingerprint of an individual can be generated.

A radioactive probe is then used to identify certain fragments of DNA that may differ between
individuals.

USES OF DNA

1. DNA collected from the crime scene would be an exact match to DNA from the culprit. In this
DNA fingerprint, all four bands from the crime scene sample match Suspect 3 an exact match.
2. DNA fingerprinting can be used to determine paternity.

A plasmid – a small circle of DNA, typically found in bacteria, that is separate from the majority of
bacterial DNA located in nucleoid. It is a vehicle for storing and studying genes.

Multiple cloning site- this site contains recognition site for specific restriction enzymes.

Origin of Replication a sequence of DNA at which replication is initiated on chromosome, plasmid or

virus/.
Antibiotic Resistance – These are bacterial extra chromosomal elements that carry genes conferring
resistance to one or more antibiotics.