151. An improved selection method for λlacZ− phages based on galactose sensitivity
- Author
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Mientjes, Edwin J., van Delft, Joost H. M., op't Hof, Brenda M., Gossen, Jan A., Vijg, Jan, Lohman, Paul H. M., and Baan, Robert A.
- Abstract
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.- Published
- 1994
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