Please cite as: CSH Protocols; 2006; doi:10.1101/pdb.prot3730

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Construction of cDNA Libraries

Stage 5: Fractionation of cDNA by Gel Filtration through Sepharose CL-4B

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 100 words of the full text of this article appear below.


INTRODUCTION

Unused linkers, adaptor-linkers, and low-molecular-weight products created by digestion of linkers in Step 4 are removed by size-exclusion chromatography.


MATERIALS

Ethanol

Optional, please see Step 5.

Marker DNA

The marker DNA should be end-labeled fragments ranging in size from 200 bp to 5 kb.

recipe caution Sodium acetate (3 M, pH 5.2)

recipe TE (pH 7.6)

recipe TE (pH 7.6) containing 0.1 M NaCl

recipe Tris-Cl (1 M, pH 8.0)

cDNA

Use the cDNA prepared in Construction of cDNA Libraries Stage 4: Attachment of Linkers or Adaptors, Steps 13 and 14.


METHOD

1. Use a hypodermic needle with a bent end to pull the cotton wool . . . [Full Text of this Article]


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