Please cite as: CSH Protocols; 2006; doi:10.1101/pdb.prot3235

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Preparation of DNA for Pulsed-field Gel Electrophoresis: Isolation of Intact DNA from Yeast

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

Yeast cells are first treated enzymatically to break down the cell walls and then resuspended in low-melting-temperature agarose plugs. The DNA is liberated by infusing the plugs with lysis buffer and proteases. This method is used to prepare both conventional and artificial yeast chromosomes.


MATERIALS

recipe EDTA (0.05 M, pH 8.0)

recipe L buffer

recipe L buffer with proteinase K and Sarkosyl

Amend the L buffer to a final concentration of 1% (w/v) Sarkosyl. Store L buffer with Sarkosyl at room temperature, and . . . [Full Text of this Article]


METHOD


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