Book chapter · Journal article
Codon optimizing for increased membrane protein production: a minimalist approach
Stockholm University1
Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark2
Bacterial Cell Factories, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark3
Research Groups, Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark4
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.
| Language: | English |
|---|---|
| Publisher: | Springer |
| Year: | 2016 |
| Pages: | 53-61 |
| Series: | Methods in Molecular Biology |
| Journal subtitle: | Methods and Protocols |
| ISBN: | 1493936352 , 1493936379 , 9781493936359 and 9781493936373 |
| ISSN: | 10643745 and 19406029 |
| Types: | Book chapter and Journal article |
| DOI: | 10.1007/978-1-4939-3637-3_4 |
| ORCIDs: | Nørholm, Morten |
