Design and total synthesis of a gene for bovine rhodopsin

In an effort to gain deeper insight into the process of visual excitation we have undertaken studies of the structure-function relationships of the visual pigment rhodopsin, retinal GTPase, rhodopsin kinase, and related proteins by the technique of site-specific mutagenesis. As the anticipated use of mutagenesis is extensive, we elected to use the approach of restriction fragment replacemenet (Lo et al. 1984). To facilitate these studies, we chemically synthesized a gene for bovine rhodopsin. The synthetic gene contains 28 unique restriction sites distributed throughout the sequence that define target fragments for mutageneisis which are, in general, no more than 60 base pairs (bp) in length. Mutations are introduced into the gene by simply replacing specific fragments with synthetic counterparts which contain the desired codon changes (Lo et al. 1984). We describe here the design, total chemical synthesis and in vitro expression of this gene.