This document provides information about DNA amplification. It begins with definitions of DNA amplification as a technique used to produce multiple copies of a target DNA sequence. It then discusses the history of DNA amplification, including that Kary Mullis developed the polymerase chain reaction (PCR) technique in the 1980s. The document outlines the principle and components of PCR, including DNA template, primers, DNA polymerase, dNTPs, and buffer solution. It describes the working of each component and the PCR procedure involving repeated heating and cooling cycles. Finally, it discusses different types of DNA amplification like emulsion PCR, bridge amplification, and DNA nanoball generation and lists some applications of DNA amplification.

1. DNA
Amplification
Minahil Khalid
Roll no 12
Mphil-MMG
2 Semester

2. Defination.
05
03
01
02 History.
Components/Working
of each component.
Working principle .
• Table of contents
04
Procedure
06
Types of DNA
Amplification.
07 Application.

3. “A technique used for the production
of multiple copies of a sequence of
target DNA”.
Definition •

4. Discovered by?
First DNA AmplificationTechnique?
Gobind Korana (recipient of the Nobel Prize in 1968)
was reported to describe a process of in vitro DNA
amplification involving oligonucleotide primers and
DNA polymerase
First Person TO Design PCR?
PCR is a technique used in the lab to make millions of
copies of a particular section of DNA
The technique was developed by Kary
Mullis in the 1980s
• History

5. Principle of DNA Amplification
❑ PCR is a DNA amplification technique
based on the principle of DNA
polymerization reaction.
❑ It relies on thermal cycling consisting of
repeated cycles of heating, cooling,
melting, and enzymatic replication of
target DNA using DNA polymerase,
primer sequence, and dNTPs.

6. Components of DNA Amplification
1. DNA template
2. Two PCR primers
3. DNA polymerase
4. Deoxynucleoside triphosphates
(dNTPs)
5. buffer solution.

7. Wor ing o each component

8. DNA Template
• DNA template is the target DNA molecule that
needs to be amplified
• The optimal quantity of DNA depends on the
composition of the DNA and DNA polymerase.
• Usually, 0.1–1 ng of plasmid DNA is sufficient for
a 50 µL PCR reaction mixture

9. dNTPS
• They are the building blocks from which the DNA
polymerase synthesizes a new DNA strand.
• dNTPs consist of four basic nucleotides - dTTP,
dCTP, dGTP, and dATP.

10. Buffer solution.
• It serves as a chemical environment to maintain
the activity and stability of the DNA polymerase.
• MgCl2 &KCL are the most commonly used buffer

11. DNA Primers
• These are short nucleotide
sequences (approximately 15–30 bases) that base
pair to a specific portion of the DNA being
replicated.
• Two sets of DNA Primer are required for the
reaction
❑ Forward Primer binds to the Template DNA
❑ Reverse Primer binds to the other complimentary
stand.

12. Taq DNA Polymerase.
• Taq DNA polymerase isolated from Thermus
aquaticus growing in hot springs acts best at 72
degrees centigrade
• DNA polymerases recognize primers as start tags
in order to start the synthesis and replication of
target DNA

13. Procedure
H-bond broken.
By heating the mixture to near boiling Cool mixture to a temp of primer
DNA-hybrid
dNTPs for replication
Warm mixture to allow replication
❑ Repeat process for 30-40 cycles.

14. Types of DNA Amplification
Three different types of
amplification used are:
❑ Emulsion PCR.
❑ Bridge amplification.
❑ DNA nanoball generation.

15. Emulsion PCR.
❖ PCR components, such as template DNA, polymerase, primers,
nucleotides, and minerals are soluble in water but not oil.
❖ This forms the basis for the concept behind emulsion PCR.
❖ It amplifies DNA molecules in physically separated water-in-oil
droplets

17. Bridge Amplification.
• DNA molecules are repeatedly replicated on a glass
flow cell containing complementary oligonucleotides
• Illumina sequencing is based on a technique called
bridge amplification.

19. DNA nanoball generation.
▪ This technique was developed by Complete Genomics.
▪ This technology does not use PCR amplification but
instead clones a library of DNA fragments into
a retroviral vector to produce circles.

20. Applications
❑ Cloning
❑ Genotyping
❑ Mutation detection
❑ Pathogen detection
❑ Forensics

21. Thank you.