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Foto del equipo PCR Digital
Responsable del servicio
Clementina Rodellar Penella
Location

Faculty of Veterinary Medicine

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Digital PCR

The digital PCR service provides the necessary equipment and infrastructure for the detection and absolute quantification of nucleic acids using the analytical technique of digital PCR (dPCR), with high precision and sensitivity.

For this purpose, this service is based on a dPCR platform: QuantStudio™ 3D Digital PCR System (QS3D CORE SYST; CHIPS- MMX V2), which works by partitioning a DNA or cDNA sample into numerous individual PCR reactions in parallel, with some of these reactions containing the target molecule (positive), while others do not contain it (negative). It allows a single molecule to be amplified at least one million times, using TaqMan® chemistry with fluorochrome-labeled probes. After PCR analysis, the fraction of negative reactions is used to generate an absolute count of the number of molecules in the sample, without the need for standards or endogenous controls.

The use of a nanofluidic chip provides a mechanism to run thousands of PCR reactions in parallel. Each well is loaded with a sample mix, master mix and TaqMan® assay reagents, and is individually analyzed for the presence (positive) or absence (negative) of an endpoint signal. To account for wells that may have received more than one molecule of the target sequence, a correction factor is applied using the Poisson model.

Advantages of digital PCR:

  • There is no need to rely on references or standards.
  • Absolute and precise copy number determination.
  • The efficiency of the reaction has less influence on the result than in qPCR.
  • High tolerance to inhibitors and ability to analyze complex mixtures.
  • Linear detection of small expression changes.

Sectors of interest:

  • Genetics and healthcare laboratories, including hospitals.
  • Laboratories in the agri-food sector.
  • Environmental analysis laboratories.
Services

This service offers the following services:

  • Absolute quantification of nucleic acids without standard curve.
  • Gene expression studies.
  • Detection and quantification of pathogens.
  • Detection of species in food.
  • Detection of GMOs.
  • Rare allele detection.
  • Generate absolute reference standards for genetic measurements.

Service delivery modalities:

  • Experiment design, set-up, chip loading, PCR, reading and analysis of results.
  • Chip loading, PCR and reading of results.
  • Self-analysis.
Equipment

Termociclador: Applied BiosystemsTM ProflexTM PCR System

Termociclador: QuantStudio™ 3D Digital PCR Instrument

Accesorios: QuantStudio™ 3D Digital PCR Chip Loader