Patent ID: 7056668

Claim:
A method for removing unincorporated oligonucleotides from a reaction mixture, the method comprising: (a) forming a mixture comprising: (i) a DNA polymerase or nucleic acid ligase; (ii) a nuclease; (iii) an upstream oligonucleotide having a 3′ portion and a 5′ portion, wherein the 3′ portion comprises a 3′ recognition group and a 3′ terminal nucleotide; and (iv) a template nucleic acid; wherein (i) and (ii) are the same enzyme or seperate enzymes; (b) digesting the 3′ portion of the upstream oligonucleotide with the nuclease; (c) extending the digested upstream oligonucleotide with the polymerase or ligating the digested upstream oligonucleotide to a downstream oligonucleotide with the ligase, wherein the extending or ligating forms a polynucleotide product; and (d) contacting the mixture with a substrate comprising binding groups that bind the 3′ recognition group, to remove unincorporated upstream pligonucleotides from the reaction mixture.