Amplification of a Bacteriophage Library

Lloyd B. Klickstein1

1 Brigham and Women's Hospital, Boston, Massachusetts
Publication Name:  Current Protocols in Molecular Biology
Unit Number:  Unit 5.10
DOI:  10.1002/0471142727.mb0510s34
Online Posting Date:  May, 2001
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Abstract

This protocol may be used for genomic DNA or cDNA libraries. A freshly packaged and titered library is adsorbed to log phase plating bacteria. The mixture is then plated at high density and allowed to grow until the plaques are just subconfluent. The phage are eluted from the plate by overnight incubation with phage buffer and the library is titered and stored.

     
 
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Table of Contents

  • Section V: Amplification of Transformed or Packaged Libraries
  • Commentary
  • Tables
     
 
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Materials

Basic Protocol 1:

  Materials
  • LB medium containing 0.2% maltose and 10 mM MgSO 4 (unit 1.1)
  • Suitable host (Table 5.10.1)
  • In vitro packaged phage library (units 5.7 & 5.8)
  • Top agarose (unit 1.1),warmed to 47°C
  • 150‐mm H plates (unit 1.1), warmed to 37°C
  • Suspension medium (SM; unit 1.11)
  • Chloroform
  • Dimethyl sulfoxide (DMSO)
  • Additional reagents and equipment for titering bacteriophage (unit 1.11)
    Table 5.0.1   MaterialsSuitable Escherichia coli Host Strains for Amplifying Lambda‐Constructed Libraries

    Vector E. coli host Relevant host genotype
    λgt10 C600hflA hflA
    λgt11 Y1088 SupF, lacIq (no anti‐biotics needed)
    EMBL 3 or 4 P2392, Q359, NM539 P2 lysogen
    Charon 4A LE392 SupF

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Figures

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Literature Cited

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