① Buffer solution with pH3.5 should be used for electrophoretic separation of four nucleotides. At this pH value, the negative charges of these four mononucleotides are very different, and they all move to the positive electrode, but the moving speed is different, and the order is UMP & gtGMP & gtAMP & gtCMP.

(2) pH8.0 should be adopted, which can make the nucleotide more negatively charged and facilitate adsorption on anion exchange resin column. Although nucleotides have more negative charges at pH 1 1.4, too high pH is not conducive to separation.

③ When the nonpolar adsorption of resin is not considered, the elution speed is determined according to the negative charge number of nucleotides, and the elution order is CMP >；; AMP & gtGMP & gtUMP in fact there is nonpolar adsorption between nucleotide and polystyrene anion exchange resin, and the nonpolar adsorption of purine base is three times that of pyrimidine base. Due to the interaction between electrostatic adsorption and nonpolar adsorption, the elution order is CMP >: >: UMP & gtGMP .