The E. coli S30 Extract for Circular DNA simplifies the transcription/translation of DNA sequences cloned in plasmid or λ vectors, providing a powerful tool for identifying and characterizing polypeptides. The investigator needs only to supply the cloned DNA containing the appropriate prokaryotic promoter and ribosome binding sites. The S30 Extract for Circular DNA Templates is prepared by modifications of the method described by Zubay from an E. coli strain B deficient in OmpT endoproteinase and lon protease activity. This results in a greater stability of expressed proteins that would otherwise be degraded by proteases if expressed in vivo. The S30 in vitro system also allows higher expression levels of proteins that are normally expressed at low levels in vivo due to the action of host-encoded repressors.
For more information, see the Protocols & Applications Guide.
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- Regulation of prokaryotic gene expression.
- Identification of cloned DNA gene products or mapping of polypeptides to defined DNA fragments.
- Rapid screening for translation from eukaryotic or prokaryotic DNAs cloned in vectors designed for high expression in E. coli.
- Screening of compounds for inhibition of bacterial translation.
- Zubay, G. (1973) Ann. Rev. Genet. 7, 267–87.
- Zubay, G. (1980) Meth. Enzymol. 65, 856–77.
- Pratt, J.M. (1984) In: Transcription and Translation, Hames, B.D. and Higgens S.J., eds., IRL Press, Oxford, 179.
- Studier, F.W. and Moffatt, B.A. (1986) J. Mol. Biol. 189, 113–30.
- Collins, J. (1979) Gene 6, 29–42.