Patent ID: 6720148
Filing Date: 2004-04-13
Classification: C07H,C12Q

Abstract:
A method of identifying a nucleotide in at least a first position in a polynucleotide sequence, comprising:providing a polynucleotide target sequence; hybridizing the target sequence with a first oligonucleotide probe, wherein: the probe comprises a first subsequence of nucleotides, a first 3â€²-terminal nucleotide, and a first florescent label coupled to the 3â€²-terminal nucleotide; the subsequence is complementary to a portion of the target sequence that is immediately adjacent to the first position; and the 3â€²-terminal nucleotide is complementary to one possible nucleotide in the first position; contacting the hybridized probe and target sequence with polymerase extension reagents in a first extension reaction mixture; measuring a level of polarized fluorescence emitted from the first extension reaction mixture, a decrease in polarized fluorescence indicating the presence of polymerase extension of the probe, the presence of polymerase extension of the probe indicating that the 3â€²-terminal nucleotide is complementary to the nucleotide in the first position; and identifying the nucleotide in the first position.