Protocol

Screening Expression Libraries Constructed in Bacteriophage Vectors

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

An expression library constructed in a bacteriophage vector is plated on an appropriate E. coli strain in the absence of isopropylthio-ß-D-galactoside (IPTG). After 2-4 hours, the plates are moved to 37°C (to stabilize any fusion proteins that are temperature sensitive), and filters impregnated with IPTG are laid on top of the developing plaques. After incubation for a further 2-4 hours, the filters, which contain imprints of the phage-encoded proteins, are removed and probed with antibody. The plates are stored at 4°C until the results of immunological screening are available.

MATERIALS

E. coli

Different bacterial host strains used in expression screening require . . . [Full Text of this Article]

TROUBLESHOOTING