Detection and identification of minor nucleotides in intact deoxyribonucleic acids by mass spectrometry

Abstract

A mass spectral method is described for the detection and identification of unusual nucleotide residues present in DNAs. Analysis by this method of intact, underivatized DNA from salmon sperm, calf thymus, mouse L-cells, wheat germ, M. lysodeikticus, E. Coli, and the bacteriophages 0X-174, fd, and lamda, yields diagnostic ions for the four common components of DNA as well as characteristic ions for 5-methyldeoxycytidine residues. The spectrum from T2 DNA contains ions indicative of 5-hydroxymethyldeoxycytidine and 5-methyldoxycytidine components but no ions corresponding to deoxycytidine residues. The DNAs of phages fd and 0X-174 also display ion products indicative of N6-methyldeoxyadenosine residues. Additional series of ions in the spectra of all four bacteriophage DNAs suggest the presence of 5-substituted deoxyuridine residues. The detection method exhibits considerable sensitivity in that amounts of DNA as low as 0.01 A260nm units can be used in the analysis, and thus, the procedure should prove of some value in the detection and location of modified components in specific regions of the various genomes by analysis of the appropriate endonuclease restriction fragments.