Critical reflections on synthetic gene design for recombinant protein expression.

Gene synthesis enables the exploitation of the degeneracy of the genetic code to boost expression of recombinant protein targets for structural studies. This has created new opportunities to obtain structural information on proteins that are normally present in low abundance. Unfortunately, synthetic gene expression occasionally leads to insoluble or misfolded proteins.

Recombinant cloning strategies for protein expression.

A variety of methods to create specific constructs for protein expression, in a broad range of organisms, are available nowadays. Restriction enzyme-free, ligation-independent and recombinase-based cloning methods have enabled high-throughput protein expression for structural and functional studies. These methods are also instrumental for modification of target genes including gene truncations, site-specific mutagenesis and domain swapping.