COMPOSITIONS, METHODS AND SYSTEMS FOR POLYMERASE CHAIN REACTION ASSAYS

摘要

The present disclosure provides methods, devices, systems and compositions for detecting nucleic acids in polymerase chain reaction assays, such as droplet digital polymerase chain reaction (ddPCR) assays. The present disclosure provides methods, devices, systems and compositions for detecting nucleic acids in ddPCR assays using intercalating dyes. A dual surfactant system with at least one fluorosurfactant and at least one non-ionic non-fluorosurfactant may be employed for droplet generation and nucleic acid detection.

主权项

1. A method of generating a plurality of droplets, comprising:
(a) providing an oil composition comprising an oil and a fluorosurfactant, wherein said fluorosurfactant is a triblock copolymer comprising polyethylene glycol (PEG) polymer covalently linked to polyhexafluoropropylene oxide (PFPE) at both ends; wherein the covalent linkage between the PEG block and PFPE blocks is by amide bonds or ester bonds at both ends, or an amide bond at one end and an ester bond at the other end; (b) providing an aqueous composition comprising a target nucleic acid, a non-ionic non-fluorosurfactant and an intercalating dye; and (c) contacting the oil composition of a) with the aqueous composition of b), thereby generating a plurality of droplets suspended in a continuous phase.