The regulatory region of the L-arabinose operon: its isolation on a 1000 base-pair fragment from DNA heteroduplexes.

A DNA fragment containing the l-arabinose operon regulatory region of Escherichia coli was purified from DNA heteroduplexes formed between opposite strands of two non-defective ara transducing phage. The phage and arabinose gene orientation is such that the heteroduplex contains two single-stranded “bubbles”. The ara regulatory region and short portions of the flanking araB and araC genes are in the short duplex between the “bubbles”. Extensive regions of homology between the phage genomes allowed nearly half of the DNA renatured from a mixture of the two phage DNAs to be in the form of heteroduplexes. Digestion of the reannealed DNA containing heteroduplexes and homoduplexes with the easily purified, single-strand specific nuclease S₁ yielded the 1000 (1017 ± 20, n = 36) base-pair ara DNA duplex plus half and whole phage-length duplexes. The larger DNA duplexes were selectively precipitated by polyethylene glycol before the final purification by preparative electrophoresis on polyacryl-amide gels. By these methods 10 to 20 μg of the 1000 base-pair DNA fragment were purified.