Alm:PCR

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(New page: Standard PCR reactions * ~100 ng template * ~100 pmole each primer * buffer to 1X * polymerase * denature 94-95°C * anneal 5°C less than lowest primer hyb temp * extend 1’/kb to be amp...)
Current revision (12:58, 3 May 2008) (view source)
 
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* anneal 5°C less than lowest primer hyb temp
* anneal 5°C less than lowest primer hyb temp
* extend 1’/kb to be amplified  
* extend 1’/kb to be amplified  
 +
 +
PCR Master Mix (2.5X)
 +
* 62.5 U/ml Taq DNA Polymerase
 +
* 125 mM KCl
 +
* 75 mM Tris-HCl, pH 8.3
 +
* 3.75 mM Mg(OAc)2
 +
* 500 uM each dNTP
 +
5'-CATTAG can be useful to add to prevent exonuclease activity from degrading recognition sites
5'-CATTAG can be useful to add to prevent exonuclease activity from degrading recognition sites
[[Category:Alm]]
[[Category:Alm]]
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[[Category:Protocols]]
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[[Category:Protocol]]

Current revision

Standard PCR reactions

  • ~100 ng template
  • ~100 pmole each primer
  • buffer to 1X
  • polymerase
  • denature 94-95°C
  • anneal 5°C less than lowest primer hyb temp
  • extend 1’/kb to be amplified

PCR Master Mix (2.5X)

  • 62.5 U/ml Taq DNA Polymerase
  • 125 mM KCl
  • 75 mM Tris-HCl, pH 8.3
  • 3.75 mM Mg(OAc)2
  • 500 uM each dNTP


5'-CATTAG can be useful to add to prevent exonuclease activity from degrading recognition sites

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