A plasmid cloning vehicle allowing a positive selection for inserted fragments

We describe a plasmid cloning vehicle, pTR262, which allows a strong positive selection (resistance to tetracycline) for transformants bearing plasmids which have DNA insertions. pTR262 is derived from plasmid pBR322 and contains the cI gene and adjacent regulator region oRpR or the bacteriophage lambda. The expression of the tetracycline resistance (tet-r) gene(s) in pTR262 requires transcription from pR and is repressed by the cI gene product, lambda repressor. Insertion of a DNA fragment into the HindIII or Bc/I sites in pTR262 inactivates the cI gene and allows expression of the tet-r gene(s) in the host bacterium. A 100-fold increase in the number of tetracycline-resistant transformants is obtained when HindIII- or Bc/I-generated fragments are added to a ligation mixture containing HindIII- or Bc/I-digested pTR 262 DNA.