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A novel in vitro transcription-translation system: accurate and efficient synthesis of single proteins from cloned DNA sequences

Stüber, Dietrich and Ibrahimi, Ibrahim and Cutler, Daniel and Dobberstein, Bernhard and Bujard, Herrmann

In: The EMBO Journal, 3 (1984), Nr. 13. pp. 3143-3148.

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Abstract

A system is described which permits the efficient synthesis of single proteins in vitro. The essential element in this expression system is a strong promoter derived from coliphage T5 which produces, with high efficiency, specific RNAs in capped or uncapped form, depending upon the experimental conditions used. The transcription-coupled capping of RNA allows the direct translation of the RNA in eukaryotic extracts from wheat germ as well as from HeLa cells. The synthesis of three different proteins is reported, including lysozyme, which is shown to be translocated across membranes when appropriate assay conditions are used. The simplicity of the experimental procedure, the high purity and specific activity of the [35S]methionine-labefled proteins produced offer a number of possibilities for the study of structure-function relationships of proteins.

Item Type: Article
Journal or Publication Title: The EMBO Journal
Volume: 3
Number: 13
Date Deposited: 02. Mar 2009 11:01
Date: 1984
Page Range: pp. 3143-3148
Faculties / Institutes: Service facilities > Center for Molecular Biology Heidelberg
Subjects: 570 Life sciences
Uncontrolled Keywords: transcription-coupled translation , T5 promoter , membrane translocation
Schriftenreihe ID: Works by Bernhard Dobberstein
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