Preparation of amino acid mixtures for cell-free expression systems

Here we present a procedure for preparing amino acid mixtures--having both the desired composition and a physiological pH--at high concentrations for cell-free expression systems. Up to 2.1 mg/mL of active protein was synthesized in batch mode reactions with an all Escherichia coli cell-free expression system. Our method is fast, easy to execute, and economically advantageous compared to expensive commercial kits, making it useful for high-throughput experiments, incorporation of nonstandard amino acids, and cell-free metabolic engineering.