[A model prokaryotic promotor. III. Cloning and study of functional activity in vivo]

The efficiency of two schemes of oligonucleotide-directed insertion mutagenesis was studied in comparison with standard cloning of DNA duplexes based on blunt-end ligation. Using new approaches, the 30-bp consensus-like prokaryotic promoter was inserted in proper orientation in a promoter-testing plasmid at an almost 100% frequency. Data on the marker gal operon expression and S1-nuclease mapping of the transcription initiation points indicate formation of an active promoter in the region of the insertion.