Abstract: Disclosed is a method for determining the nucleotide sequence of a target oligonucleotide. In this method a single-stranded ligation product is prepared which contains a target oligonucleotide-to-be sequenced and an auxiliary oligonucleotide. A primer complementary to a portion of the auxiliary oligonucleotide and having a label covalently attached thereto is annealed to the auxiliary oligonucleotide portion of the ligation product. The primer is extended with chain-extending nucleoside triphosphates and chain-terminating nucleoside triphosphates in the presence of a polymerase to yield a plurality of primer extension products. These extension products are then separated on the basis of their base length; and the nucleoside sequence of the target oligonucleotide is determined from the mobilities of the primer extension products obtained during their separation.