✦ LIBER ✦

High efficiency cell-free synthesis of proteins: Refinement of the coupled transcription/translation system

✍ Scribed by Wieslaw Kudlicki; Gisela Kramer; Boyd Hardesty

Publisher: Elsevier Science
Year: 1992
Tongue: English
Weight: 734 KB
Volume: 206
Category: Article
ISSN: 0003-2697
DOI: 10.1016/0003-2697(92)90383-i

No coin nor oath required. For personal study only.

✦ Synopsis

Two modifications are introduced to convert the Escherichia coli cell-free extract ("S30") into a high efficiency system for coupled transcription/translation of exogenously added genes. (a) The ribosome fraction collected from the S30 by ultracentrifugation is used. It contains all the proteins necessary for gene expression but has lost the vast majority of soluble proteins that might interfere with purification and enzymatic activity of product formed. (b) Plasmids containing coding sequences to be expressed are not linearized thus enhancing their stability by avoiding their degradation. These two modifications not only improve protein synthesis in a static system but allow gene expression over 20-40 h in the continuous-flow cell-free system. Both prokaryotic and eukaryotic proteins have been synthesized in this system.

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