Patent ID: 8133672

Claim:
A method for nucleotide sequencing comprising: a) providing a reaction mixture having: (i) a polymerase enzyme, (ii) polymerase reaction conditions including cofactors, and (iii) polymerase reaction substrates including a primed template and nucleotides, such that a reaction comprising incorporation of the nucleotides into a growing nucleic acid occurs; and b) observing the reaction mixture to determine the incorporation of nucleotides into the growing nucleic acid; wherein the polymerase enzyme, the polymerase reaction conditions, and the polymerase reaction substrates are selected such that the reaction exhibits two kinetically observable steps, each of which kinetically observable steps proceeds from an intermediate in which a nucleotide or a polyphosphate product is bound to the polymerase enzyme or each of which kinetically observable steps proceeds from an intermediate in which the nucleotide and the polyphosphate product are not bound to the polymerase enzyme, and c) using the observed incorporation of nucleotides to determine a sequence of the template.