Processes of Genetic Engineering enhancement of a present trait by increasing

or disrupting the expression of the desired

gene.
What is Genetic Engineering?
• It has an application in the pharmaceutical,
- The wide variety of plant-based products and
industrial, agricultural, medical, and other
fortified formulas in the market today has
industries.
been made possible by artificial selection of
desired genes through genetic engineering.

Genetic Engineering - a gene modification process

wherein the Deoxyribonucleic acid (DNA) is
transferred from one organism to another.

GMO’s (Genetically Modified Organisms)

• Genetically modified organisms have been
subject for public scrutiny whether it is safe to
use or ethically accepted.
Example of Genetic Engineering:
• These challenge the researchers to prove the
significance of GMOs as a breakthrough in
science.

Classical Breeding vs. Genetic Engineering

Processes of Genetic Engineering

 Stage 1. DNA Cleavage
 Stage 2. Production of Recombinant DNA
 Stage 3. Cloning
 Stage 4. Screening
Classical breeding focuses on the mating of organisms
with desirable qualities or traits while genetic Processes:
engineering involves molecular techniques to modify
1. DNA Cleavage 
the traits of a target organism.
- a restriction endonuclease is used to cleave
Example of Genetic Engineering: the source DNA into a different set of
fragments. 

• The modification of traits may involve the

introduction of new traits into an organism or
 2. Production of Recombinant DNA 
- The fragments of DNA are inserted into
plasmids that have been cleaved with the
same restriction endonuclease as the source
DNA.

Methods of Introducing Plasmids to Host

Organism
 1. Biolistic
 2. Plasmid insertion by Heat Shock
Treatment
 3. Electroporation
Plasmid insertion
Methods:

1. Biolistic
- This technique uses a “gene gun” to fire DNA-
coated pellets on plant tissues. Cells that are
able to survive and take up the expression
plasmid coated pellets can acquire the ability
to express the designed protein.
3. Cloning
- The plasmids serve as vectors that can
introduce the DNA fragments into cells---
usually, but not always bacteria. As each cell
produces, it forms a clone of cells that all
contain the fragment-bearing vector.

4. Screening
- The clones containing a specific DNA fragment 2. Plasmid insertion by Heat Shock Treatment
of interest are identified from the clone - is a process used to transfer plasmid DNA into
library. bacteria. The target cells undergo a
pretreatment procedure to increase the pore
sizes of their plasma membranes.

- The pretreatment using Calcium chloride
makes the cells “competent” for the
introduction of the plasmid DNA, then the
cells are incubated with the desired plasmid
at about 4°C for about 30 minutes.
- During this time, the plasmids concentrate
near the cells. Afterward, 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. The rapid rise and drop of
temperature increase and decrease the pore
sizes in the membrane, respectively.
Recombinant Cell
- These are cells that are made by combining
genetic material from two different sources.
 1. Selection of plasmid DNA containing cells
 2. Selection of transformed cells with the
desired gene
 3. Polymerase Chain Reaction (PCR) detection
of plasmid DNA
Methods:
1. Selection of plasmid DNA containing cells
- A selection marker within the inserted
plasmid DNA sequence allows the selection of
“transformants”. Usually, an antibiotic
resistance gene is included in the plasmid
DNA. This mechanism allows only
“transformed” cells to survive in the presence
of the antibiotic (e.g. ampicillin).
2. Selection of transformed cells with the
desired gene
- The most general procedure for screening
clone libraries to find a particular gene is
hybridization - the cloned genes form base-
pairs with complementary sequences on
another nucleic acid.
- The complementary nucleic acid is called as
probe because it is used to probe for the
presence of the gene of interest.
3. Polymerase Chain Reaction (PCR) detection
of plasmid DNA
- the presence of the desired gene in the
inserted plasmids may also be confirmed
through PCR amplification.
- PCR reactions specific for the desired gene
may be done using DNA from cells that would
confirm the presence of the gene within the
samples.
PCR reactions specific for plasmid sequences will
confirm/identify the type of plasmid used for the
transformation through the following steps:
• Step 1. Denaturation.
• Step 2. Annealing of Primers.
• Step 3. Elongation.