Saturation mutagenesis is a random mutagenesis technique, in which a single codon or set of codons is randomised to produce all possible amino acids at the position.
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Method
Saturation mutagenesis is commonly achieved by artificial gene synthesis, with a mixture of nucleotides used at the codons to be randomised. Different degenerate codons can be used to encode sets of amino acids. Because some amino acids are encoded by more codons than others, the exact ratio of amino acids cannot be equal. Additionally, it is usual to use degenerate codons that minimise stop codons (which are generally not desired). Consequently, the fully randomised 'NNN' is not ideal, and alternative, more restricted degenerate codons are used. 'NNK' and 'NNS' have the benefit of encoding all 20 amino acids, but still encode a stop codon 3% of the time. Alternative codons such as 'NDT', 'DBK' avoid stop codons entirely, and encode a minimal set of amino acids that still encompas all the main biophisical types (anionic, cationic, aliphatic hydrophobic, aromatic hydrophobic, hydrophilic, small).
Applications
Saturation mutagenesis is commonly used to generate variants for directed evolution.