Codon Compression Algorithms for Saturation Mutagenesis

Saturation mutagenesis is employed in protein engineering
and genome-editing efforts to generate libraries that span amino acid
design space. Traditionally, this is accomplished by using degenerate/
compressed codons such as NNK (N = A/C/G/T, K = G/T), which
covers all amino acids and one stop codon. These solutions suffer from
two types of redundancy: (a) different codons for the same amino acid
lead to bias, and (b) wild type amino acid is included within the library.
These redundancies increase library size and downstream screening efforts.
Here, we present a dynamic approach to compress codons for any desired
list of amino acids, taking into account codon usage. This results in a
unique codon collection for every amino acid to be mutated, with the
desired redundancy level. Finally, we demonstrate that this approach can
be used to design precise oligo libraries amendable to recombineering and
CRISPR-based genome editing to obtain a diverse population with high
efficiency.