Alm:PCR: Difference between revisions
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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...) |
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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]] | ||
[[Category: | [[Category:Protocol]] | ||
Latest revision as of 10:58, 3 May 2008
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
