Endy:Site-directed Mutagenesis

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==Primer Design==
==Primer Design==
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*See [[Designing Primers | this page]] for general advice on primer design.
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*See [[Designing Primers | this page]] for general advice and software suggestions for primer design.
*Primers should contain the mutations to be introduced but should anneal to the sequence to be mutated.  You will need a forward and reverse primer.
*Primers should contain the mutations to be introduced but should anneal to the sequence to be mutated.  You will need a forward and reverse primer.
*Primers should be between 25 and 45 base-pairs in length.
*Primers should be between 25 and 45 base-pairs in length.

Revision as of 10:09, 2 July 2005

Primer Design

  • See this page for general advice and software suggestions for primer design.
  • Primers should contain the mutations to be introduced but should anneal to the sequence to be mutated. You will need a forward and reverse primer.
  • Primers should be between 25 and 45 base-pairs in length.
  • The mutation(s) should be in the middle of the primers.
  • Mutation efficiency is greatly influenced by the purity of the primers. Its worth getting purified primers. No need for 5' phosphorylation.
  • See the Stratagene manual for more detailed information.

Experimental Protocol

  • See the Stratagene manual for the suggested protocols.
  • Comments and usage notes on this protocol will be added when the protocol has been tested.
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