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. Author manuscript; available in PMC: 2020 Jul 30.
Published in final edited form as: J Vis Exp. 2019 Jun 20;(148):10.3791/59729. doi: 10.3791/59729

Figure 2. The DE3 cassette/pET vector dual system explained.

Figure 2

(A) The sketched genome of pET vector transformed BL21(DE3) pLysS E. coli bacteria. The native bacterial genome carries a DE3 cassette (see panel B), as well as a lac gene that constantly expresses lac repressor units. The non-native pET vector carries the protein gene inserted between a T7 polymerase promoter and terminator sequence. More details in panel B. (B) The DE3 cassette of the native bacterial genome encodes the information for T7 polymerase in terms of an E. coli RNA polymerase operon. This protein, however, is not expressed because the lac repressor unit prevents the RNA polymerase protein from binding. Hence no T7 polymerase is expressed and no exogenous protein is expressed. (C) Application of the chemical IPTG (Table of Materials) distorts the structure of lac repressor units and prevents them from binding to the DE3 cassette. As a result, RNA polymerase can now bind to the cassette, for which T7 polymerase is expressed, as is exogenous protein eventually. Please click here to view a larger version of this figure.