KLE SOCIETY’S S.
NIJALINGAPPA
COLLEGE RAJAJIAGAR-
560010
DEPARTMENT OF ZOOLOGY
SEMINAR TOPIC: POLYMERASE
CHAIN REACTION
FROM: Thoshitha V TO: Dr. Tejaswini. V.
II Year BtZ Nandi
U18EB21S0454 HoD Zoology
WHAT IS PCR?
PCR or Polymerase Chain Reaction
is a technique used in molecular
biology to create several copies of
a certain DNA segment.
This technique was developed in
1983 by Kary Mullis, an American
biochemist.
PCR has made it possible to
generate millions of copies of a
small segment of DNA.
This tool is commonly used in the
COMPONENTS OF PCR
Components Of PCR constitutes the following:
• DNA Template– The DNA of interest from the
sample.
• DNA Polymerase– Taq Polymerase is used. It is
thermostable and does not denature at very
high temperatures.
• Oligonucleotide Primers- These are the short
stretches of single-stranded DNA
complementary to the 3’ ends of sense and
anti-sense strands.
• Deoxyribonucleotide triphosphate–
These provide energy for polymerization
and are the building blocks for the
synthesis of DNA. These are single units
of bases.
• Buffer System– Magnesium and
Potassium provide optimum conditions
for DNA denaturation and renaturation.
It is also important for fidelity,
polymerase activity, and stability.
STEPS OF
PCR
The PCR involves three major cyclic
reactions:
1. Denaturation
2. Annealing
3. Elongation
1. Denaturation
• The first step in PCR is denaturation.
Denaturation is required to separate the
double-stranded DNA sample.
• It is done at 94-98 ℃ for 20-30 seconds. It
breaks the hydrogen bonds present
between base pairs.
• Denaturation leads to the formation of
single strands of DNA.
• The single strands now act as a template for
the production of new strands of DNA.
• The temperature should be provided for a
longer time to ensure the separation of the
two strands.
2. Annealing
• The second step is the annealing of the
primer.
• Here, the primers bind to their
complementary sequences on the template
DNA.
• Primers are single-strand sequences of DNA
or RNA around 20 to 30 bases in length.
• They serve as the starting point for the
synthesis of DNA.
• The two separated strands run in the
opposite direction and consequently there
3. Elongation
• At this step, the temperature is raised to
72-80℃. The bases are added to the 3’ end
of the primer by the Taq polymerase
enzyme.
• This elongates the DNA in the 5’ to 3’
direction. The DNA polymerase adds about
1000bp/minute under optimum conditions.
• Taq Polymerase can tolerate very high
temperatures. It attaches to the primer and
adds DNA bases to the single strand. As a
result, a double-stranded DNA molecule is
obtained.
• These three steps are repeated 20-40 times
in order to obtain a number of sequences of
DNA of interest in a very short time period
Applications of PCR
The following are the applications of PCR :
Medicine
Testing of genetic disease mutations.
Monitoring the gene in gene therapy.
Detecting disease-causing genes in the
parents.
Forensic Science
Used as a tool in genetic fingerprinting.
Identifying the criminal from millions of
people.
Paternity tests
RT-PCR is commonly used to detect the
SARS-CoV-2 viral genome. It is a variant of
PCR and uses the enzyme reverse
transcriptase to produce DNA
complementary to the viral RNA.
Research and Genetics
Compare the genome of two organisms in
genomic studies.
In the phylogenetic analysis of DNA from
any source such as fossils.
Analysis of gene expression.
Gene Mapping
THANK-YOU