Lidstrom:QuikChange Site-Directed Mutagenesis

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Back to [[Lidstrom:Protocols|Protocols]]
Back to [[Lidstrom:Protocols|Protocols]]
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==QuikChange Basics ==
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* You can buy a kit from Agilent or [[Richard_Lab:Site_Directed_Mutagenesis|use your own ingredients]].
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* The marketing term "lightning" refers to an improved Dpn1, that degrades template (unmutated template) more efficiently.
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== Two (main) QuikChange kits are available ==
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* QuikChange Lightning.
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** For mutations contained within a single primer
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* QuikChange Lightning Multi
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** For mutations contained in >1 primer
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** Can be used for mutations contained in 1 primer, but this kit is more expensive than the single kit and likely has lower efficiency.
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* Both kits:
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** come with chemically competent XL10-Gold.  This is perhaps the most valuable part of the kit.
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== Tips ==
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* Though the lightning Dpn1 enzyme claims to be effective in 5 minutes, you might as well incubate it for 30 minutes, just in case it lowers your unmutated background further.

Revision as of 08:55, 12 June 2014

Back to Protocols

QuikChange Basics

  • You can buy a kit from Agilent or use your own ingredients.
  • The marketing term "lightning" refers to an improved Dpn1, that degrades template (unmutated template) more efficiently.

Two (main) QuikChange kits are available

  • QuikChange Lightning.
    • For mutations contained within a single primer
  • QuikChange Lightning Multi
    • For mutations contained in >1 primer
    • Can be used for mutations contained in 1 primer, but this kit is more expensive than the single kit and likely has lower efficiency.
  • Both kits:
    • come with chemically competent XL10-Gold. This is perhaps the most valuable part of the kit.

Tips

  • Though the lightning Dpn1 enzyme claims to be effective in 5 minutes, you might as well incubate it for 30 minutes, just in case it lowers your unmutated background further.
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