Hybrid P L tl Promoter with Dual Regulation Control

Abstract

The newly constructed P_Ltl hybrid promoter is composed of the operator and promoter sequences of tac and the −35 to −135 region of the phage λ P_L promoter, which contains the AT-rich block and the OL2 and OL3 segments. Transcriptional properties of P_Ltl were examined and compared with tac and λ P_L as reference promoters. The hybrid P_Ltl exhibits different and improved properties over tac promoter in four ways: (i) when repressed, the repression is almost complete; (ii) after complete induction, the hybrid P_Ltl promoter shows a 1.4–2 times higher expression; (iii) the hybrid P_Ltl promoter permits flexible gene expression because it can be utilized under either or both repression controls simultaneously; (iv) the P_Ltl promoter permits enhanced expression of genes encoding products with unknown properties. When compared with the strong promoter P_L from phage λ, results with the P_Ltl promoter in lacZ fusion constructs show higher levels of β-galactosidase activity. We also constructed a plasmid vector, pP_Ltl7G, which contains the hybrid P_Ltl promoter with a polylinker sequence at its 3′ end, which facilitates efficient fusions of foreign genes in any of the reading frame.