In vitro gene fusions that join an enzymatically active beta-galactosidase segment to amino-terminal fragments of exogenous proteins: Escherichia coli plasmid vectors for the detection and cloning of translational initiation signals

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Abstract
The construction and use of a series of plasmid vectors suitable for the detection and cloning of translational control signals and 5'' coding sequences of exogenously derived genes are reported. In these plasmids, the 1st 8 codons of the amino-terminal end of the lactose operon .beta.-galactosidase gene, lacZ, were removed, and unique BamHI, EcoRI and SmaI (XmaI) endonuclease cleavage sites were incorporated adjacent to the 8th codon of lacZ. Introduction of DNA fragments containing appropriate regulatory signals and 5'' coding sequences into such lac fusion plasmids led to the production of hybrid proteins consisting of the carboxyl-terminal segment of a .beta.-galactosidase remnant plus a peptide fragment that contained the amino-terminal amino acids encoded by the exogenous DNA sequence. These hybrid peptides retained .beta.-galactosidase enzymatic activity and yielded a Lac+ phenotype. Such hybrid proteins are useful for purifying peptide sequences encoded by exogenous DNA fragments and for studies relating the structure and function of specific peptide segments.