Determining the ratio of prototype nucleotides to predetermined nucleotides, which is preferably greater than 1:1, by doping in walk-through mutagenesis

摘要

A method of walk-through mutagenesis of a nucleic acid encoding a prototype polypeptide of interest is described. The method comprising selecting a predetermined amino acid and one or more target regions of the polypeptide, and synthesizing a mixture of oligonucleotides containing at each sequence position in the target region, either a prototype nucleotide that is required for synthesis of the prototype amino acid of the polypeptide, or a predetermined nucleotide that is required for synthesis of the predetermined amino acid, in which during the synthesis, the ratio of available prototype nucleotides, to available predetermined nucleotides, is greater than 1:1.