Klenow Fragment

It is a large fragment of DNA polymerase I enzyme. It results from the proteolytic cleavage, of DNA polymerase I, caused by the protease subtilisin. Of the 3 main purposes of DNA polymerase I, the Klenow fragment possesses two of them, which are:

·         5’ → 3’ polymerase activity.

·         3’ → 5’ exonuclease activity.

So basically, the Klenow fragment is a DNA polymerase I enzyme, without the 5’ → 3’ exonuclease activity. But having those two properties makes the Klenow fragment have applications such as:

®     The preparation of radioactive DNA probes.

®     DNA sequencing through the Sanger method.

®     Synthesis of second strand of cDNA.

Reference:

Sambrook, J., Fritsch, E.F. and Maniatis, T. (1989). Molecular Cloning: A Laboratory Manual. (2^nd ed.), 5.40-5.43. Cold Spring Harbor: Cold Spring Harbor Laboratory Press.