Medical Microbiology

410 7 GeneralVirology

PolymeraseChainReaction

Separation of the double strand by application

of heat

Primer- ( ) dependent synthesis using Taq

polymerase ( )

Products of original DNA

(= 1 st PCR product)

• defined 5 ́ end


• undefined 3' end


• present from 1st cycle;
  linear increase
  Products of 1st PCR product
  (= 2 nd PCR product)


• defined 5 ́ and 3 ́ ends


• present from 2nd cycle;
  linear increase
  Products of 2nd PCR product
  (= 3 rd and subsequent PCR products)


• defined 5 ́ and 3 ́ ends


• present from 3rd cycle;
  exponential increase

5'

3'

3'

5'

5' 3'

5' 5'

3' 5'

5'

3'

3'

5' 5'

3'

3'

5'

5'

3'

3'

5'

a

3' 5'

5' 3'

3'

b

3' 5'

5' 3'

5'

5' 3'

Fig. 7. 9 aTwooligonucleotideprimersarehybridizedtotheDNAdoublestrands,

whichhavebeenseparatedbyheating.Aheat-stablepolymerase(e.g.,Taqpoly-

merasefromThermusaquaticus)isthenaddedandextendstheseprimersalong

thelengthof,andcomplementaryto,thematrixstrand.Theresultingdouble

strandsarethenonceagainseparatedbyheatandthereactionisrepeated.

bTheDNAstrandsproducedinthefirstcycle(1stgeneration)haveadefined 50 end

(correspondingtotheprimer)andanundefined 30 end.Allofthesubsequent

daughterstrands(2ndtonthgeneration)haveauniform,definedlength.

7

Kayser, Medical Microbiology © 2005 Thieme