Promega Corporation

Novagen, Inc.

Emd Biosciences, Inc.

Thomas R Van Oosbree

Robert C Mierendorf

David V Thompson

Mark McCormick

Gregory S Beckler

Yuping Ambuel

John F Van Herwynen

There is now a market for kits containing the necessary components for performing protein synthesis reactions using template DNA of the experimenter. One category of existing system for performing protein synthesis reactions is based on an S-30 extract from the bacteria. It is disclosed here that a simple fractionation process can dramatically improve the performance of an S-30 prokaryotic protein synthesis reaction mixture. In one embodiment, the fractionation is a simple freezing and thawing of an S-30 extract combined with a supplemental mix, followed by centrifugation. The resulting fractionated S-30 reaction mixture yields more full-length target protein and less non-full length or non-target protein than possible using prior art S-30 systems.

E. coli extract for protein synthesis

A method for performing coupled in vitro transcription and translation reactions is disclosed. In the transcription reaction, the quantity of mRNA produced is limited to a level of less than about 2.5 micrograms in a 10 microliter volume prior to the translation elements being added. Limiting the level of mRNA produced prevents saturation of the translational processes and thus aids in the efficiency and fidelity of the translation process.

Method for in vitro protein synthesis

A method for performing coupled in vitro transcription and translation reactions is disclosed In the transcription reaction, the quantity of mRNA produced is limited to a level of less than about 2.5 micrograms in a 10 microliter volume prior to the translation elements being added. Limiting the level of mRNA produced prevents saturation of the translational processes and thus aids in the efficiency and fidelity of the translation process.

A method for coupling transcription and translation from DNA using a solution comprising a eukaryotic cell-free extract comprising adding a sufficient amount of a magnesium compound to the extract to raise the magnesium concentration to a level where RNA is transcribed from DNA and RNA translates into protein.

Growing community of inventors

Madison, WI, United States of America

Thomas R Van Oosbree