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Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis.

ABSTRACT: The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40-45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were changed in a single experiment. This method is especially useful for codon optimization, where doublet or triplet rare codons can be changed using a single mutagenic primer set, in a single experiment.

SUBMITTER: Hussain H

PROVIDER: S-EPMC5138438 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Combined Overlap Extension PCR Method for Improved Site Directed Mutagenesis.

Hussain Hasnain H Chong Nikson Fatt-Ming NF

BioMed research international 20161122

The combined overlap extension PCR (COE-PCR) method developed in this work combines the strengths of the overlap extension PCR (OE-PCR) method with the speed and ease of the asymmetrical overlap extension (AOE-PCR) method. This combined method allows up to 6 base pairs to be mutated at a time and requires a total of 40-45 PCR cycles. A total of eight mutagenesis experiments were successfully carried out, with each experiment mutating between two to six base pairs. Up to four adjacent codons were ...[more]

PMID: 27995143

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