Left-handed DNA. Cloning, characterization, and instability of inserts containing different lengths of (dC-dG) in Escherichia coli

Abstract

Recombinant pBR322 derivatives were constructed containing tracts of (dC-dG) sequences which are 58, 32, 26, and 10 base pairs (bp) in length in conjunction with a 95-bp fragment containing the Escherichia coli lac operator-promoter. The biological properties of these plasmids were unusual since deletions in the (dC-dG) regions, but not in the pBR322 nor the lac segments, were frequently observed; segments of (dC-dG) longer than approximately 50 bp were not stable but suffered deletions. Segments of approximately 30 bp or shorter were stable in most cases. The (dC-dG) tracts seemed to enhance recA-mediated recombination when they were of suitable length and were cloned into certain sites in the recombinants. Also the (dC-dG)-containing plasmids were less supercoiled (by 6-12 turns) than expected, relative to control plasmids, after isolation from E. coli hosts. These recombinant plasmids were used in the following paper to evaluate the properties of segments containing the unorthodox left-handed conformations.