Presented by: Maryam iqbal

Roll. No (57)
Presented to: Mam Fazeelat
Outlines:
1. Theory
2. What is PCR
3. Discovery of PCR
4. Principle of PCR
5. Components of PCR
6. Steps of PCR
7. Applications of PCR
8. Types of PCR
9. Role of PCR In Detecting Corona Virus
 Theory:
Polymerase chain reaction, better known as PCR, is one of the technology that
not only made impact on scientific community but also effect many aspects of lives.
More than 30 years ago, the introduction of recombinant DNA technology as a tool
for the biological science.
Molecular cloning allowed the study of individual genes of living organism, but this
technique was dependent on obtaining a relatively large quantity of pure DNA.
Researchers found it very difficult and laborious for obtaining large quantity of
DNA.
 What is PCR
• The polymerase chain reaction is a fast and inexpensive
technique used to amplify “copy” small segment of DNA
rapidly and accurately to make million or even billions of
copies of DNA in a very short time.
• Mane made version of DNA replication
• In vitro technique ( in laboratory )
• Sometimes called “ molecular photocopying “
• PCR is the most important inventions of the 20 th century in
molecular biology.
 Discovery of PCR

• The idea of PCR is credited to Kary Mullis, a

research scientist at California Biotech
Company, in 1983.
• For this work Mullis received the Noble
prize in Chemistry along with his
colleagues.
• Nobel prize in 1993
 Principles of PCR

1. Purpose
2. Components
3. Steps
 Purpose of PCR

 Purpose of PCR Technique in vitro ( Test tube ) is

amplification of specific DNA sequences
 To make several million copies in just a few hours.
PCR Components
 Template strand:

 Also known as antisense strand

 It is a parent strand on
which
DNA polymerase

act and mRNA, is synthesized

 Its direction from 3” to
5”
 Primer in PCR

 It is a short single stranded

nucleic acid
 They contain the
complementary sequence to
DNA sequences
 Two primers are required in
PCR
 Nucleotides:
 Four nucleotides are
required in PCR
 Building blocks of
DNA
Taq polymerase
 Like DNA replication in an organism, PCR
required a DNA polymerase enzyme that
makes new strands of DNA using existing
strand as template
 The DNA polymerase used in PCR is Taq
polymerase
 It is an enzyme come from specie of
bacteria
 Found in hot spring
 It is a highly thermostable
Magnesium ions:

• Mg2+ have variety of effect such as

• Act as a co-factor for Taq polymerase
• Help in binding property of primer with
the template
 Steps of PCR
1.Denaturation
2.Annealing
3.Extension or elongation
 Denaturation:
 Melting or separation of
DNA strands
 Temperature 94-95°C
 At high temperature the
hydrogen bonds break
 20-30 seconds
 Annealing
Temperature lowered to
50-60 degree
20-40 seconds
Attachment of primers
with the single stranded
template
Extension or elongation:
In this step DNA polymerase
synthesize a new DNA strand
complimentary to the template
DNA
The temperature at this step
depend on the DNA
polymerase used such as
Taq polymerase
Cycle according to
Temperature range
Complete cycle:
Applications of PCR
Diagnosis of genetic diseases
Amplification of microbial DNA
Checking contamination
Estimation of gene expression
PCR cloning
Food pathogen detection using PCR
Types of PCR
• Real-Time PCR 
• Reverse-Transcriptase (RT-PCR)
• Multiplex PCR.
• Nested PCR.
• High Fidelity PCR.
• Fast PCR.
• Hot Start PCR.
• GC-Rich PCR.
Detection of coronavirus

• RNA virus
• Real time RT–PCR is one of the most widely used laboratory methods
for detecting the COVID-19 virus.
• In order for a virus like the COVID-19 virus to be detected early in
the body using real time RT–PCR, scientists need to convert the RNA
to DNA.
• This is a process called ‘reverse transcription’.
• They do this because only DNA can be copied — or amplified —
which is a key part of the real time RT–PCR process for detecting
viruses.