Identification of Nucleic Acids and Oligonucleotides by Capillary Zone Electrophoresis with the Four Marker Technique

Abstract

The paper describes the improvement brought by the four marker technique to the repeatability of the separation of macromolecules [nicotinamide nucleotides (NADPH and NADH), one adenine compound (ATP), two oligonucleotides (135-oligomer and 84-primer containing 135 and 20 bases, respectively) and N2-5′-deoxyquanosine-benzo[a]pyrene-diolepoxide (dG5′BaDE)] by capillary zone electrophoresis with UV detection at the wavelength of 220 nm. Optimized conditions [3-(cyclohexylamino)-1-propanesulphonic acid as the organic electrolyte solution with pH of 11.2, and concentration of 50 mM] were used for the separation. The counter-ion in the system was sodium. The macromolecules could be separated within 20 minutes. The four marker compounds were (-)mandelic acid, meso-2,3-diphenylsuccinic acid, 1,2-phenylenediacetic acid and o-phthalic acid, which were used for the calculations of the mobilities for the analytes. The identification of the macromolecules was confirmed with the coefficients of identification. The reliability of the identification was increased up to 75-fold relative to the use of the absolute migration times.