Screening for highly active plasmid promoters via fusion to beta-galactosidase gene

A plasmid containing promoter-deleted inactive beta-galactosidase gene [1] was used to select promoters of the pEP 121 plasmid [2]. Colonies of cells harboring reactivated beta-galactosidase gene were identified by their red color on McConkey plates. The quantitative amounts of beta-galactosidase produced in each clone were estimated by assaying enzyme activity and by measuring the specific beta-galactosidase protein following fractionation of total cells' proteins on polyacrylamide gel. A wide range of enzyme activities was observed. The most active promoter isolated was shown to promote beta-galactosidase production more efficiently, compared with the original beta-galactosidase promoter, amounting to 20% of all cell proteins. Such highly active promoters may be utilized in the future, to promote expression of cloned genes in bacteria.