| 摘要 |
The present invention provides a droplet actuator device and methods for multiplexed PCR amplification and detection of target amplicons within a single droplet. The methods of the invention combine quantitative real-time PCR (qPCR) amplification with fluorescence-based sequence specific detection technologies for amplified DNA. In one embodiment, fluorescently-labeled oligonucleotide probes may be used for hybridization-based multiplexed detection of target amplicons. The methods of the invention generally involve combining the necessary reactants to form a PCR-ready droplet and thermal cycling the droplet at temperatures sufficient to result in amplification of one or more target nucleic acids. Fluorescence-based detection techniques may be used for end-point or real-time analysis of DNA amplification. For end-point analysis, the accumulation of a signal, e.g., a fluorescence signal, is measured after the amplification of the target sequence is complete. For real-time analysis, the signal is measured while the amplification reaction is in progress. |
