title: mRNA stability and plasmid copy number effects on gene expression from an inducible promoter system.
abstract: The effects of mRNA stability and plasmid copy number on gene expression in Escherichia coli were evaluated by constructing multicopy (pMB1-based) and low-copy (F-based) plasmids containing an arabinose-inducible promoter system, the lacZ reporter gene, and mRNA-stabilizing 5' hairpin structures. Product formation and cell growth were evaluated under a number of inducer concentrations. The introduction of a 5' hairpin into the untranslated region of the mRNA resulted in significantly higher gene expression from the multicopy plasmids at low inducer concentrations and increased gene expression from the low-copy plasmids across all inducer concentrations investigated. With high inducer concentrations, expression from high-copy plasmids significantly slowed cell growth, whereas expression from the low-copy plasmids had little effect on growth rate. At inducer concentrations between 1 x 10(-4) and 4 x 10(-4)%, the productivity of low-copy plasmids containing the 5'-hairpin was equal to or greater than that from multicopy plasmids. Together, these two gene expression strategies may find important use in metabolic engineering and heterologous gene expression.
authors: Department of Chemical Engineering, University of California, Berkeley, California 94720-1462, USA.
authors: T
authors: T
authors: Carrier
authors: K L
authors: KL
authors: Jones
authors: J D
authors: JD
authors: Keasling
mesh_terms: D001089:Arabinose / Q000378:metabolism; D015152:Blotting, Northern; D002455:Cell Division; D015870:Gene Expression; D005821:Genetic Techniques; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000502:physiology*; D013997:Time Factors; D001616:beta-Galactosidase / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D001089:Arabinose; D001616:beta-Galactosidase