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US 7,229,799 B2Granted
System and method for nucleic acid sequencing by polymerase synthesis
AssigneeLI-COR, Inc.
InventorJohn G. K. Williams
Life of this patent
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Abstract
This invention relates to improved methods for sequencing and genotyping nucleic acid in a single molecule configuration. The method involves single molecule detection of fluorescent labeled PPi moieties released from NTPs as a polymerase extension product is created.
Claims19
A method for genotyping or sequencing a single target nucleic acid molecule, said method comprising:
- i) immobilizing onto a solid support a target nucleic acid or a nucleic acid polymerase to form a solid support comprising more than one site or location each bearing only one single individual molecule of polymerase or target nucleic acid;
- ii) contacting the solid support with the target nucleic acid when the polymerase is immobilized or the polymerase when the target nucleic acid is immobilized, a primer nucleic acid which complements a region of the target nucleic acid, at least two types of fluorescently labeled nucleotide triphosphates (NTPs), wherein each type of NTP has a different fluorescently label on the γ-phosphate;
- and
- iii) determining the incorporation of a fluorescently labeled NTP into a single molecule of the primer stand, by detecting the fluorescently labeled γ-phosphate of the labeled NTP, which is then cleaved away from the fluorescently labeled NTP, thereby genotyping or sequencing said single target nucleic acid.
1 more element
A method for genotyping or sequencing a single target nucleic acid molecule, said method comprising:
- i) conducting a nucleic acid polymerase reaction in the presence of a single target nucleic acid, a primer, a nucleic acid polymerase and a plurality of fluorescently labeled nucleotide triphosphates (NTPs), the primer hybridizing to said target nucleic acid, wherein each type of nucleotide triphosphate (NTP) is differently fluorescently labeled on the γ-phosphate , and wherein said nucleic acid polymerase reaction results in the production of a fluorescently labeled phosphate;
- and
- ii) detecting each type of said fluorescently labeled phosphate individually to genotype or sequence said single target nucleic acid molecule.
Dependencies are inferred from the claim text.