A general method for maximizing the expression of a cloned gene.

Abstract
A method is presented which utilizes a combination of restriction endonuclease cleavage and digestion with Escherichia coli exonuclease III and Aspergillus oryzae nuclease S1, that allows the positioning of a restriction fragment bearing the promoter of the lacZ gene of E. coli at virtually any distance in front of any cloned gene. This method was used to examine the effect on protein production of gene-promoter separation for the cro gene of phage .lambda. and to produce plasmids that, upon transformation into appropriate E. coli hosts, direct the synthesis of up to 190,000 cro protein monomers per cell.