Michael R. Green; Joseph Sambrook
Cold Spring Harb. Protoc., 2020
https://doi.org/10.1101/pdb.top100743

Abstract

Escherichia coli DNA Pol I can carry out three enzymatic reactions: It possesses 5′ → 3′ DNA polymerase activity and 3′ → 5′ and 5′ → 3′ exonuclease activity. Pol I can be cleaved by mild treatment with subtilisin into two fragments; the larger fragment is known as the Klenow fragment. The Klenow fragment retains the polymerizing activity and the 3′ → 5′ exonuclease of the holo-enzyme but lacks its powerful 5′ → 3′ exonuclease activity. These enzymes and their applications in molecular cloning are introduced here.