PCR AND ITS TYPES

CONTENTS

• PCR Definition

• PCR history

• PCR procedure

• PCR types

• PCR applications

• Conclusion

• References

• POLYMERASE CHAIN REACTION

Pcr

PCR STEPS

PCR STEPS

PCR STEPS

Cycling conditions for amplifying longer PCR products

Step Time/cycles Temperature

Initial activation step 2 min 95°C

3-step cycling

Denaturation 10 s 94°C

Annealing 1 min 50–68°C*

Extension 1 min/kb

Number of cycles 40 cycles 68°C

End of PCR cycling Indefinite 4°C

PCR

PCR TYPES

PCR TYPE

• Overlap-extension PCR or Splicing by overlap extension (SOEing) :

• Genetic engineering technique

• used to splice together two or more DNA fragments OR complementary sequences.

• It is the technique enables creation of specific and long DNA constructs.

• It can also introduce deletions, insertions or point mutations into a DNA sequence.

• Nested PCR

• This PCR increases the sensitivity

• Two sets of primers,

• A double process of amplification .

• The first set of primers allow a first amplification. The product of this PCR is subjected to a second PCR using the second set of primers.

• Primers used in the second PCR are specific to an internal amplified sequence in the first PCR. specificity of the first PCR product is verified with the second one.

• Semi quantitative PCR

• An approximation to the relative amount of nucleic acids present in a sample,

• The markers commonly used are

• Apo A1 and B actin.

• Amplification product is separated by electrophoresis

• Multiplex PCR

• Multiplex PCR is an adaptation of PCR which allows simultaneous amplification of many

sequences.

• This technique is used for diagnosis of different diseases

• Multiplex PCR can detect different pathogens in a single sample.

PCR TYPES

Applications of PCR and impact on science

• PCR and its different variations are highlighted as the most commonly used in laboratories and research institutes.

• Thus, these have contributed to identification

• characterization of several organisms and understanding of physiopathology of diverse

• diseases in human, animal and plants.

• MEDICINE

• identification of microorganisms

assurance of blood

• Forensic

• As a basic procedure to investigate Deaths

• (paternity testing)

• Evidence from minimal samples of saliva, semen or other tissue debris

AGRICULTURE

• As conventional PCR or qPCR have also facilitated research in

• Detection of pathogens in plants, animals, and the environment; understanding of their

• Epidemiology and, development of new diagnostic tests, treatments or vaccines.

• Selective DNA isolation• Isolation of DNA fragments from genomic DNA

by selective amplification of a specific region of DNA.

• This use of PCR augments many methods, such as generating hybridization Probes and DNA cloning which require larger amounts of DNA, representing a specific DNA region.

• PCR supplies these techniques with high amounts of pure DNA, enabling analysis of DNA samples Possible.