Patent ID: 6136543
Filing Date: 2000-10-24
Classification: C12Q

Abstract:
A method determining a base sequence of a template DNA comprising the steps of:(a) hybridizing a primer with a template DNA;(b) performing a complementary strand extension reaction using a polymerase for extending said hybridized primer or an extended primer produced by repeating said step (b) to following step (d), by incorporating a single chemically modified nucleotide of four kinds of chemically modified nucleotides to 3'-terminus of said hybridized primer or said extended primer, in the presence of said four kinds of chemically modified nucleotides, wherein said single chemically modified nucleotide is complementary with a base sequence of said template DNA, and each of said four kinds of chemically modified nucleotides has a chemical modification for preventing a continuous progress of said complementary strand extension reaction, after incorporating said single chemically modified nucleotide to 3'-terminus of said hybridized primer or said extended primer;(c) detecting said single chemically modified nucleotide incorporated to 3'-terminus of said hybridized primer or said extended primer, and determining a kind of a base of said single chemically modified nucleotide incorporated to 3'-terminus of said hybridized primer or said extended primer;(d) carrying out a chemical reaction for changing a chemical structure of said single chemically modification nucleotide incorporated to 3'-terminus of said hybridized primer or said extended primer, in order to bring about a state at which said complementary strand extension reaction for extending said hybridized primer or said extended primer can proceed; and(e) repeating said step (b) to said step (d), and determining said base sequence of said template DNA one base by one base sequentially based on the kinds of bases of said chemically modified nucleotides incorporated to said extended primer by said complementary strand extension reaction.