DE3containing

DE3-containing refers to Escherichia coli strains that carry the DE3 lysogen, a defective lambda prophage encoding the T7 RNA polymerase. In these strains, the gene for T7 RNA polymerase is placed under the control of the lacUV5 promoter, enabling IPTG-inducible production of T7 RNAP and, consequently, high-level transcription from T7 promoters on plasmids used for recombinant protein expression.

Mechanism and use: When IPTG is added, the lac repressor is inactivated, T7 RNA polymerase is produced, and transcription from plasmid-borne T7 promoters is initiated. The resulting messenger RNA drives strong expression of the target gene. This system is widely used with pET- and related vectors, offering robust, tightly regulatable expression suitable for many proteins.

Common strains: The canonical host is BL21(DE3). Derivatives such as BL21(DE3) pLysS/pLysE and other DE3-containing strains like C41(DE3) and C43(DE3) are used to improve expression of toxic or problematic proteins by providing additional regulation or control of T7 RNAP activity, for example through T7 lysozyme to dampen basal transcription.

Practical considerations: While DE3-containing strains enable high-level expression, they can exhibit basal expression or toxicity for some proteins. This can be mitigated by using lower IPTG concentrations, autoinduction media, modifying growth conditions, or employing strains that modulate T7 RNAP activity. Choice of plasmid copy number and promoter strength also influences outcomes.

History: The DE3 system was developed in the late 1980s by Studier and colleagues as part of the T7 expression system, revolutionizing recombinant protein production in E. coli.

See also: T7 expression system, pET system, lac operon, IPTG induction.