Patent ID: 8309308

Claim:
A method for determining the amount of template nucleic acid present in a sample comprising the steps of: i) bringing into association with the sample all the components necessary for nucleic acid amplification, and all the components necessary for a bioluminescence assay for nucleic acid amplification including: a) a nucleic acid polymerase, b) the substrates for the nucleic acid polymerase, c) at least two primers, d) a thermostable luciferase, e) luciferin, f) an enzyme that converts PPi to ATP, wherein the enzyme is not ATP sulphurylase, and any other required substrates or cofactors of the enzyme of part f), wherein the substrate or cofactor is not PPi, wherein when the enzyme of part f) is pyruvate orthophosphate dikinase, the substrates added in part g) are phosphoenolpyruvate and AMP; and subsequently: ii) performing a nucleic acid amplification reaction of the template nucleic acid, wherein generated copies are themselves further copied; iii) coupling the production of PPi to light output from the bioluminescence assay by converting generated PPi to ATP and using luciferase to follow the changes in the concentration of ATP by monitoring the intensity of light output from the bioluminescence reaction; and iv) determining the amount of template nucleic acid present in the sample.