An improved selection method for λlacZ− phages based on galactose sensitivity
The determination of thelacZ mutant frequency in λgt10lacZ phage vectors isolated from the transgenic mouse strain 40.6 (MutaMouse), requires the screening of large numbers of phages on β-galactosidase activity. Existing methods rely on distinguishing a few white plaques on X-gal containing plates amongst a multide of blue ones which is both time-consuming and expensive. The new screening method described here employs the galactose sensitiveEscherichia coli C lacZ recA galE strain into which a multicopy plasmid has been introduced, which results in over-expression of thegalK andgalT genes. In the presence of phenyl-β-d-galactopyranoside, a substrate for β-galactosidase, this leads to the suppression of λlacZ+ phage propagation without affecting the ability of λlacZ− phages to form plaques. With this method it is possible to screen 1.5×106 phages on a single 9-cm Petri dish. Furthermore, the need for blue/white screening has been eliminated.
Key words40.6 transgenic mouse MutaMouse mutant selection λgt10lacZ phage galE
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