Construction of a DNA-polymerase I overproducing plasmid and isolation of the enzyme.

The polA gene of Escherichia coli coding for DNA polymerase I was cloned under the control of bacteriophage lambda promoter pL and gene N in a high copy number plasmid vector. The chromosomally located lambda cIts repressor gene kept the synthesis of the polA gene product at 28 degrees C at a low level. Raising the temperature to 43 degrees C resulted in inactivation of the repressor and overproduction of DNA polymerase I, which could easily be purified to homogeneity.