Plasmid pDOF2 was generated at Calagen Biosciences, LLC for robust expression of toxic proteins. Gene of interest (GOI) is expressed from the phage T7 promoter. To shut off the basal level expression of the GOI, two tandem copies of the lac operator (lacO) sequence were inserted downstream of the T7 promoter (T7/O2), and the wild-type lacI inhibitor is expressed from the lacIq promoter.
Calagen Biosciences also has two E. coli strains to use with the pDOF2 plasmid (and related plasmids) for toxic protein expression. BL21T7L is a BL21 derivative that has the T7 RNA polymerase gene expressed from Plac/O2 (a lacUV5 promoter containing two lac operators downstream), and BL21T7A is a BL21 derivative that has the T7 RNA polymerase gene expressed from the tightly-regulated, arabinose-inducible PBAD promoter. The T7pol cassettes are inserted at the lambda phage attB site.