Labelled nucleotides

Abstract

Nucleosides and nucleotides are disclosed that are linked to detectable labels via a cleavable linker group.

1 .- 25 . (canceled) 
     
     
         26 . A method for determining the sequence of a target single stranded polynucleotide, comprising:
 a) providing four different types of nucleotides, wherein each of the nucleotides comprises: a 3′-hydroxyl group and a base that is linked to a detectable label via a cleavable linker; wherein the detectable label and the cleavable linker block the incorporation of a further nucleotide into a growing polynucleotide strand, wherein the detectable label linked to each nucleotide can be distinguished upon detection from the detectable label used for the other three nucleotides;   b) incorporating one nucleotide from step a) into a complementary strand of the target single stranded polynucleotide;   c) removing non-incorporated nucleotides;   d) detecting the detectable label of the incorporated nucleotide of step b), thereby determining the type of nucleotide incorporated; and   e) removing the detectable label of the incorporated nucleotide of step b).   
     
     
         27 . The method of  claim 26 , further comprising: repeating steps a)-e) one or more times, thereby determining the sequence of the target single stranded polynucleotide. 
     
     
         28 . The method of  claim 26 , wherein the target single-stranded polynucleotide is immobilized on a solid support. 
     
     
         29 . The method of  claim 26 , wherein each of the nucleotides for incorporation is a deoxyribonucleotide triphosphate. 
     
     
         30 . The method of  claim 29 , wherein the base is a purine or a pyrimidine. 
     
     
         31 . The method of  claim 30 , wherein the base is a deazapurine. 
     
     
         32 . The method of  claim 26 , wherein the detectable label is a fluorophore. 
     
     
         33 . The method of  claim 26 , wherein the linker is an acid labile linker or a photolabile linker. 
     
     
         34 . The method of  claim 33 , wherein the linker comprises an azido moiety or a disulfide linkage. 
     
     
         35 . The method of  claim 26 , wherein the incorporation of the nucleotide is accomplished by a terminal polymerase, a terminal transferase, or a reverse transcriptase. 
     
     
         36 . The method of  claim 35 , wherein the incorporation of the nucleotides is accomplished by a terminal polymerase. 
     
     
         37 . A method of labelling a nucleic acid molecule, the method comprising incorporating a nucleotide into the nucleic acid molecule, wherein the nucleotide comprises: a 3′-hydroxyl group and a base that is linked to a detectable label via a cleavable linker; wherein the detectable label and the cleavable linker block the incorporation of a further nucleotide into the nucleic acid molecule. 
     
     
         38 . A kit comprising:
 a) four individual nucleotides, wherein each nucleotide comprises a 3′-hydroxyl group and a base that is linked to a detectable label via a cleavable linker, wherein the detectable label and the cleavable linker block the incorporation of a further nucleotide into a growing polynucleotide strand, and wherein the detectable label linked to each nucleotide can be distinguished upon detection from the detectable label used for the other three nucleotides; and   b) packaging materials therefor.   
     
     
         39 . The kit of  claim 38 , wherein two or more of the individual nucleotides are mixed. 
     
     
         40 . The kit of  claim 38 , wherein each of the nucleotides is a deoxyribonucleotide triphosphate. 
     
     
         41 . The kit of  claim 40 , wherein the base is purine, deazapurine, or pyrimidine. 
     
     
         42 . The kit of  claim 38 , wherein the detectable label is a fluorophore. 
     
     
         43 . The kit of  claim 38 , wherein the linker is an acid labile linker or a photolabile linker. 
     
     
         44 . The kit of  claim 43 , wherein the linker comprises an azido moiety or a disulfide linkage. 
     
     
         45 . The kit of  claim 38 , further comprising a polymerase and a buffer appropriate for the action of the polymerase.