Analysis of Phosphorothioated Oligo DNA (VN-50 2D)

A good separation of phosphorothioate-oligomers was achieved without the use of ion-pair reagents under the HILIC mode using Shodex HILICpak VN-50 2D. The gradient mode developed used 50 mM ammonium formate (pH9.5) and acetonitrile.

Sample: 1 µL
Synthesized phosphothioated oligo DNA 20 mer (crude)

(A*T*A*C*C*G*A*T*T*A*A*G*C*G*A*A*G*T*T*T ; * means phosphorothioated position)

1.0 mg/mL (in H₂O)

Column       : Shodex HILICpak VN-50 2D (2.0 mm I.D. x 150 mm)
Eluent       : (A); 50 mM HCOONH4 aq. (pH9.5)/(B); CH3CN
               Linear gradient ;
               (B %) 64 to 58 % (0 to 10 min), 58 % (10 to 15 min),
                    58 to 64 % (15 to 15.01 min), 64 % (15.01 to 20 min)
Flow rate    : 0.25 mL/min
Detector     : UV (260 nm) (small cell volume)
Column temp. : 40 °C