New reagent for discrimination of single- and double-stranded regions in DNA

It was found that DNA alkylation at the N-7 guanine with the bulky alkylating reagent, N,N,N'-tri-(β-chloroethyl)-N-(p-formylpheny)propylene diamine-1,3 (TFP) is much diminished when DNA is double-stranded. We report here an application of this reaction for probing the hairpin structure in the palindrome-containing single-stranded (ss) DNA fragment of 377 bases prepared from the EcoRI-BamHI fragment of plasmid pBR322. 5'-Labeled ss fragment was modified with TFP and cleaved by piperidine hydrolysis at the alkylated guanine residues according to the Maxam-Gilbert procedure. Guanines in the hairpin formed by palindrome of 9 bp were protected from TFP action, while dimethyl sulfate modified all guanines.