BACTERIAL ACID-PHOSPHATASE GENE FUSIONS USEFUL AS TARGETS FOR CLONING-DEPENDENT INSERTIONAL INACTIVATION

The Morganella morganii phoC gene, encoding a class A acid phosphatase , was used to generate gene fusions with modified amino-terminal moiet ies of the Escherichia coli lacZ gene carrying a multiple-cloning site flanked by phage-specific promoters and recognition sites for univers al sequencing primers. The corresponding hybrid proteins retained a Ph oC-like enzymatic activity which is easily detectable by a plate histo chemical assay, rendering similar gene fusions potentially useful as t argets for cloning-dependent insertional inactivation. Cloning experim ents performed in plasmids carrying similar lacZ-phoC fusions confirme d their usefulness as cloning vectors for direct screening of recombin ants. As compared to conventional lacZ alpha-complementation-based vec tors, which can only be used in E. coli hosts carrying specific lacZ m utations, the lacZ-phoC fusion-based vectors can be used in combinatio n with any E. coli host and require a less expensive histochemical ass ay for screening of recombinants, while retaining all the advantageous features that made the former so popular as general purpose cloning v ehicles.