Open AccessReaction conditions affect the specificity of bromoacetaidehyde as a probe for DNA cruciforms and B—Z junctions
Author(s) -
Michael J. McLean,
Jacquelynn E. Larson,
Franz Wohlrab,
Robert D. Wells
Publication year - 1987
Publication title -
nucleic acids research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 9.008
H-Index - 537
eISSN - 1362-4954
pISSN - 0305-1048
DOI - 10.1093/nar/15.17.6917
Subject(s) - dna supercoil , cruciform , holliday junction , biology , dna , pbr322 , plasmid , biophysics , reactivity (psychology) , ionic strength , dna replication , biochemistry , chemistry , dna repair , materials science , aqueous solution , composite material , medicine , alternative medicine , pathology
The reaction of bromoacetaldehyde (BAA) was investigated further with recombinant plasmids containing tracts of (CG)16, in pRW756, or (CA)32, in pRW777, which adopt left-handed Z-structures under the influence of negative supercoiling. The cruciform structures adopted by the inverted repeat sequences near the replication origins of the pBR322 vectors served as internal controls for the number of unpaired bases. The extent of reaction with the B-Z junctions and the cruciforms was dependent on the reaction and analysis conditions, the method of preparation of BAA, ionic conditions, and the amount of negative supercoiling. In contrast to the previous results of Kang and Wells, B-Z junctions in addition to cruciforms do react with BAA. However, more forcing conditions are required to detect this reaction since B-Z junctions appear to be less reactive than the single stranded loops of cruciforms. The site of reaction with DNA was readily mapped with high precision at the nucleotide level. Also, a simple method is described for determining the concentration of BAA as well as its intrinsic reactivity in a given ionic medium.