[21] β-Galactosidase gene fusions for analyzing gene expression in Escherichia coli and yeast

Gene fusions can be constructed either in vivo, using spontaneous nonhomologous recombination or semi-site specific transposon recombination, or in vitro, with recombinant DNA technology. This chapter describes in vitro methods and lists some recently developed β-galactosidase gene fusion vectors. With these methods, gene-controlling elements from any source can be fused to the β-galactosidase structural gene and examined in the prokaryote bacterium Escherichia coli or the lower eukaryote yeast Saccharomyces cerevisiae. β-galactosidase gene fusions can be constructed both with transcription initiation control signals and with transcription plus translation initiation control signals. The β-galactosidase gene is convenient for making translational fusions because it is possible to remove its translation initiation region along with up to at least the first 27 amino acid codons without affecting β-galactosidase enzymic activity. The focus here is on these transcription-translation fusions because they provide all the gene initiation signals from the other gene. β-Galactosidase expression from a gene fusion can be used not only to measure gene expression and regulation, but also to isolate mutations and additional gene fusions.