Bio-Rad QX200 ddPCR system
Digital PCR is a breakthrough technology that provides ultrasensitive and absolute nucleic acid quantification. This technique is particularly useful for low abundance targets, targets in complex backgrounds, allelic variants (SNPs) and for monitoring of subtle changes in target levels.
In the Droplet Digital™ PCR (ddPCR™) System, each PCR sample is partitioned into a large number of microscopic droplets prior to amplification. Each droplet is an individual PCR reaction. After end-point amplification, fluorescence is detected in the droplets in which target sequence was amplified and scored as positive. Droplets without the target sequence show little or no fluorescence and are scored as negative. Using the Poisson distribution law, the fraction of positive droplets is converted to the number of molecules in the starting sample, without the need for standard curves (absolute quantification).
The CCR Genomics Core maintains a Bio-Rad QX200 ddPCR system. This system includes a droplet generator, a C1000 Touch thermal cycler, and a droplet reader.
The QuantaSoft Analysis Pro software used for data analysis is available at the Core and free to download. To report target amplification, EvaGreen, or specific probes labeled with fluorophores FAM and/or HEX (or VIC), can be used. Multiplexing is accomplished with the two-color detection feature of the QX200 ddPCR system. To get started on ddPCR, below are some core resources:ddPCR EvaGreen TemplateddPCR Sample form Probes ddPCR User Guide