A cloning vector employing a versatile ß-glucosidase as an indicator for recombinant clones.

ABSTRACT

A mutant glucosidase, cpGluT, with activity toward chromogenic substrates (X-gal [5-bromo-4-chloro-3-idolyl-ß-d-galactoside] and indican) and a fluorogenic 4-methylumbeliferyl-ß-d-glucopyranoside (MUG) was constructed by replacing the monomeric ß-glucosidase region (E314-N326) with designed multiple cloning sites. When expressed in hosts (lacZ+ and lacZ-), a vector containing the cpGluT produced a colored or fluorescent phenotype according to the substrate supplemented on LB plates without any inducer. cpGluT is readily incorporable into customized vectors and does not require special hosts to detect recombinant plasmids, thereby making screening recombinants more effective and less expensive.