主权项 |
1. A method for sequencing a nucleic acid by detecting the identity of a nucleotide analogue incorporated into primer extension strand in a polymerase reaction, comprising:
a) providing i) sequencing primer having 3′ and 5′ ends, comprising a minor groove binder (MGB) molecule linked to the 5′ end; (ii) single-stranded template attached to a solid surface; (iii) polymerase and iv) one or more different nucleotide analogues, wherein each different nucleotide analogue comprises a base selected from the group consisting of adenine, guanine, cytosine, thymine, and uracil, and their analogues; and a unique label attached through a cleavable linker to the base or to an analogue of the base; a deoxyribose; and a cleavable chemical group at the 3′-position of the deoxyribose; 2) hybridizing the sequencing primer to the template; 3) extending the sequencing primer by incorporating a first nucleotide analogue therein with said polymerase in a polymerase reaction so as to create a primer extension strand, wherein the incorporated nucleotide analogue terminates the polymerase reaction; and 4) detecting the unique label attached to the nucleotide analogue that has been incorporated into the growing strand of DNA, so as to thereby identify the incorporated nucleotide analogue. |