Protocol

Isolation of High-molecular-weight DNA from Mammalian Cells Using Proteinase K and Phenol

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

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INTRODUCTION

This is the method of choice when large amounts of mammalian DNA are required, for example, for Southern blotting (Rapid Isolation of Mammalian DNA, Rapid Isolation of Yeast DNA, Southern Blotting: Capillary Transfer of DNA to Membranes) or for construction of genomic libraries in bacteriophage vectors. Approximately 200 µg of mammalian DNA, 100-150 kb in length, is obtained from 5 x 10⁷ cultured aneuploid mammalian cells (e.g., HeLa cells). The usual yield of DNA from 20 ml of normal blood is approx. 250 µg.

The initial stages of the procedure vary, depending on the type of . . . [Full Text of this Article]

MATERIALS

METHOD