In vitro synthesis of expression products, particularly proteins, in a cell-free system, comprises using a repetitive batch method, with periodic removal of metabolites and supplementation

摘要

Preparative in vitro synthesis of an expression product (I) in a cell-free transcription and translation system (A). Preparative in vitro synthesis of an expression product (I) in a cell-free transcription and translation system (A) comprises: (A) filling a reaction vessel with a solution containing components of (A) for a selected (I), amino acids and metabolic components that provide energy and are required for synthesis of (I); (B) performing synthesis of (I) for a predetermined time (T), without separation of substances formed and without addition of consumed synthesis substrates; (C) after expiry of T, low molecular weight metabolic products and/or reaction inhibitors are removed from solution; (D) immediately before, after or during step (c), the synthesis components that have been consumed are replaced; and (E) steps (b)-(d) are repeated, at least once, and in the last cycle, steps (c) and (d) may be omitted. An independent claim is also included for a similar method in which the reaction vessel may also include a system (A') for synthesis of a second, different product (I'), plus amino acids and required metabolic components; step (b) is as above; step (c) is omitted and step (d) is similar but includes addition of (A'), if not already present, and the final step is synthesis of (I') for a predetermined time, with separation of substances formed or addition of consumed synthesis substrates.