Patent ID: 6972174
Filing Date: 2005-12-06
Classification: C12Q

Abstract:
1. A method for identifying a target nucleotide in a nucleic acid molecule having a 3′ terminus, and a 5′ terminus with the target nucleotide located between the 3′ terminus and the 5′ terminus of the nucleic acid molecule, the method comprising: (a) mixing an oligonucleotide primer with the nucleic acid molecule in a plurality of reaction containers, wherein the oligonucleotide primer having a free 3′ hydroxyl moiety becomes hybridized to a complementary sequence on the nucleic acid molecule directly adjacent to the target nucleotide on the 3′ side; (b) providing a corresponding chain-terminating nucleotide to each reaction container, wherein the corresponding chain-terminating nucleotide is complementary to an adjacent nucleotide that is directly next to the target nucleotide on the 5′ side; (c) adding a distinct chain-extending nucleotide to each reaction container, wherein each reaction container has the same corresponding chain-terminating nucleotide but the distinct chain-extending nucleotide is different in each reaction container; (d) conducting a template dependant extension of the oligonucleotide primer by adding a polymerase reaction mixture to each of the reaction containers to give an extended primer; (e) detecting an incorporation of nucleotides into the extended primer, wherein the identity of the target nucleotide is determined from the extended primer size;