Integrated microfluidic and solid state pyrosequencing systems

摘要

The invention provides for sequencing a nucleic acid molecule based on the detection of base incorporation by the release of pyrophosphate (PPi) using a new enzyme system comprising adenosine diphosphate (ADP)-glucose pyrophosphorylase (AGPase) and its substrate ADP-glucose.

主权项

1. A method for sequencing a target nucleic acid molecule, the method comprising:
(a) contacting the target nucleic acid molecule with a pyrophosphate sequencing reagent, the pyrophosphate sequencing reagent comprising:
(i) a nucleotide triphosphate,(ii) a polymerase,(iii) a pyrophosphate to ATP converting enzyme comprising adenosine diphosphate (ADP)-glucose pyrophosphorylase and a substrate ADP-glucose; and(iv) an ATP detecting enzyme; and (b) detecting a resulting optical signal wherein the optical signal is indicative of a reaction of the pyrophosphate sequencing reagent with the target nucleic acid molecule, thereby sequencing the nucleic acid molecule.