DIGITAL PCR (EMULSION PCR)

Technology: Digital PCR is a new approach to nucleic acid detection and quantification. It offers a different method for absolute quantification and rare allele detection relative to conventional real-time quantitative PCR. Digital PCR works by partitioning a sample into many (up to 20 000) individual real-time PCR reactions; some portion of these reactions contain the target molecule (positive) while others do not (negative).

Following PCR analysis, the fraction of negative reactions is used to generate an absolute count of the number of target molecules in the sample, without reference to standards or endogenous controls. This makes ddPCR a much more sensitive solution compared to the classical qPCR reaction.

Sample partitioning allows sensitive, specific detection of single template molecules and precise quantification while mitigating the effects of target competition, making PCR amplification less sensitive to inhibition and greatly improving the discriminatory capacity of assays that differ by only a single nucleotide. Digital PCR offers the benefits over other PCR methods of absolute quantification and greatly enhanced sensitivity and dynamic range.

Services:

Genomic Laboratory of the DNA Research Center offers experiment performance on the most recent digital PCR system - QX200TM Digital Droplet PCR (BioRad) using:
EvaGreen or
TaqMan probes

Reporter: FAM, VIC or HEX
Quencher: BHQ

Applications	Advantages of Digital PCR
· Rare allele detection · Absolute quantification of gene expression and small RNA analysis · Absolute quantification of viral load · Absolute quantification of nucleic acid standards · Absolute quantification of next-generation sequencing libraries · CNV analysis · Pathogen detection · Single cell analysis	· No need to rely on references or standards · Desired precision can be achieved by increasing the number of PCR replicates · Highly tolerant to inhibitors · Capable of analyzing complex mixtures · Provides a linear response to the number of copies presentto help enable analysis of small fold-change differences

Service step by step:

1. Assistance in experiment planning.
2. Performance of ddPCR experiment.
3. Result analysis.