Poly(ethylene glycol)-oligodeoxyribonucleotide block copolymers for affinity capillary electrophoretic separation of single-stranded DNAs with a single-base difference

An affinity capillary electrophoretic method was developed to detect a single-base difference of single-stranded DNA (ssDNA). Poly(ethylene glycol)-oligodeoxyribonucleotide block copolymers (PEG-b-ODN) were prepared for use as a novel affinity ligand. We introduced a running buffer solution of PEG-b-ODN into a capillary tube, and electrophoretically separated a mixture of chemically synthesized 20 mer ssDNA (normal ssDNA) and a single-base-substituted 20 mer ssDNA (mutant ssDNA). When the base sequence of PEG-b-ODN was designed to be complementary to part of the normal ssDNA, the migration rate of the normal ssDNA was significantly decreased by reversible hybridization with PEG-b-ODN, depending on the base number of PEG-b-ODN, the salt concentration of the running buffer, and the capillary temperature. In contrast, the mobility of mutant ssDNA did not change because the interaction with PEG-b-ODN was negligible. Optimization of the analytical conditions gave two distinct peaks, one for normal and the other for mutant ssDNA, on the electropherogram, allowing for facile discrimination of the single-base difference. The results indicate that PEG-b-ODN is a promising affinity ligand for the capillary electrophoretic separation of normal and single-base mutated ssDNA.