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Polymerase Chain Reaction

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Polymerase Chain Reaction is a biochemical

technique for in vitroDNA amplification.

PCR was first described by Kary Mullis in 1983..

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. a process by which DNA is artificially

rep ca e y repea e cyc es o an n v roDNA polymerase reactions.

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.

There are three major steps in a PCR, which.

. .

.

3. extension 72 C .

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Target DNA Sequence

53 Denaturation

35

Annealing

53

35

53 Extension

5 353

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The thermal cycler (PCR machine).

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The gel electrophoresis appartus

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DNA electrophorogram

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PCR components.

DNA template.

ne or morepr mers.

DNA polymerase (e.g. Taq polymerase).

Deoxynucleotide triphosphates.

.

Divalent cations(Mg+2

).

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UsedtoidentifyapersonbycomparinghisorherDNA

withthe

DNA

in

a

iven

sam le.

patternof

DNA

fragments

generated

by

restriction

' '

.

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Electrophoresis of PCR-amplified DNA fragments.

Genetic relationships can be determined bycomparing DNA fingerprints.

This technique can also be used to determine

evolutionar relationshi s between or anisms.

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Targetgenescanbeamplifiedthensequencedto

detectmutations

(hereditary).

.

sequencecanbeanalyzedbyPCR.

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Cloning genes

,

genefromoneorganismandtheninsertingitinto

.

PCRisusedtoamplifyDNAfragments(genes).

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Archeological analysis of fossil.

PCR,to

analyze

fossil

DNA.

oldDNA

(mammoth).

The

analysis

of

Egyptian

mummies

.

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polymorphismexists.

Singlenucleotidepolymorphism.

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uan a vevers onso areuse omon orgeneexpression.

mRNAisisolatedandreversetranscribedintocDNA.

Reversetranscriptase

PCR

(RT

PCR).

RealtimePCR (RTPCR)

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amplify,isolateoridentifyaknownsequencefrom.

reversetranscriptase.

Iswidelyusedinexpressionmapping,determining.

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RealtimePCRisthetechniqueofcollectingdata

throughoutthe

PCR

process

as

it

occurs,

thus

combiningamplificationanddetectionintoasingle

step.

Thismethodusesfluorescentd esand robesto

measuretheamountofamplifiedproductinreal

time.

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