Codon optimizing for increased membrane protein production: a minimalist approach

K. Mirzadeh, S. Toddo, Morten Nørholm, Daniel O. Daley

Research output: Chapter in Book/Report/Conference proceedingBook chapterResearchpeer-review


Reengineering a gene with synonymous codons is a popular approach for increasing production levels of recombinant proteins. Here we present a minimalist alternative to this method, which samples synonymous codons only at the second and third positions rather than the entire coding sequence. As demonstrated with two membrane-embedded transporters in Escherichia coli, the method was more effective than optimizing the entire coding sequence. The method we present is PCR based and requires three simple steps: (1) the design of two PCR primers, one of which is degenerate; (2) the amplification of a minilibrary by PCR; and (3) screening for high-expressing clones.
Original languageEnglish
Title of host publicationHeterologous Expression of Membrane Proteins : Methods and Protocols
EditorsIsabelle Mus-Veteau
Number of pages9
Publication date2016
ISBN (Print)978-1-4939-3635-9
ISBN (Electronic)978-1-4939-3637-3
Publication statusPublished - 2016
SeriesMethods in Molecular Biology


  • Membrane protein
  • Protein expression
  • Codon optimization
  • Synonymous codon


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