Cite as: Cold Spring Harb. Protoc.; 2006; doi:10.1101/pdb.prot3551

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Labeling 3' Termini of Double-stranded DNA Using the Klenow Fragment of E. coli DNA Polymerase I

Joseph Sambrook and David W. Russell

This protocol was adapted from Molecular Cloning, 3rd edition, by Joseph Sambrook and David W. Russell. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, USA, 2001

The first 15% of the full text of this article appears below.


INTRODUCTION

Templates for the end-filling reaction are produced by digestion of DNA with a restriction enzyme that creates a recessed 3'-hydroxyl terminus. The Klenow enzyme is then used to catalyze the incorporation of one or more [{alpha}-32P]dNTPs into the recessed 3' terminus in a template-dependent fashion.


MATERIALS

recipe caution Ammonium acetate, 10 M

Optional, please see Step 6.

Appropriate . . . [Full Text of this Article]


METHOD


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