Please cite as: CSH Protocols; 2006; doi:10.1101/pdb.prot3929

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Ligating Plasmid and Target DNAs in Low-melting-temperature Agarose

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

Ligation in low-melting-temperature agarose is much less efficient than ligation with purified DNA in free solution and requires a large amount of DNA ligase. The method is used chiefly for rapid subcloning of segments of DNA in dephosphorylated vectors and assembling recombinant constructs.


MATERIALS

recipe 2x Bacteriophage T4 DNA ligase mixture

Restriction endonucleases

Please see Step 4.

Plasmid DNA . . . [Full Text of this Article]


METHOD


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