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Related Concept Videos

PCR01:32

PCR

Overview
Real Time RT-PCR02:57

Real Time RT-PCR

Real-time reverse transcription-polymerase chain reaction, or Real-time RT-PCR, is an analytical tool used to determine the expression level of target genes. The method involves converting mRNA to complementary DNA with the help of an enzyme known as reverse transcriptase, followed by the PCR amplification of the cDNA. These two processes can be performed simultaneously in a single tube or separately as a two-step reaction.
The real-time quantification of the number of amplified products is...
PCR - Polymerase Chain Reaction01:32

PCR - Polymerase Chain Reaction

Overview

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Fabricating Reactive Surfaces with Brush-like and Crosslinked Films of Azlactone-Functionalized Block Co-Polymers
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Interaction of quantitative PCR components with polymeric surfaces.

Asensio Gonzalez1, Ronan Grimes, Edmond J Walsh

  • 1Stokes Research Institute, University of Limerick, Limerick, Ireland.

Biomedical Microdevices
|December 21, 2006
PubMed
Summary

Polymer tubing can inhibit polymerase chain reaction (PCR) amplification by adsorbing DNA and dyes. Longer tubing or reduced sample volumes increase this inhibition, crucial for microfluidic device design.

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Area of Science:

  • Biochemistry
  • Materials Science
  • Analytical Chemistry

Background:

  • Polymerase chain reaction (PCR) is a fundamental molecular biology technique.
  • Microfluidic devices offer miniaturized platforms for biological assays.
  • Component adsorption to tubing walls can affect assay performance.

Purpose of the Study:

  • To investigate the impact of polymeric capillary tubing on PCR amplification.
  • To determine the role of tubing material, length, contact time, and flow rate on PCR inhibition.
  • To identify the specific PCR components adsorbing to tubing walls.

Main Methods:

  • Exposing PCR mixtures to various polymeric capillary tubing materials and lengths.
  • Varying contact times and flow rates during exposure.
  • Quantifying PCR inhibition and component adsorption (DNA, Sybr Green I).

Main Results:

  • No PCR inhibition observed with short tubing (40 cm) and long residence times (45 min).
  • PCR inhibition occurred with longer tubing (3 m) or reduced sample volumes, correlating with volume-to-length ratio.
  • Significant adsorption of DNA and Sybr Green I to tubing walls was detected.
  • Flow velocity did not impact PCR yield.

Conclusions:

  • PCR inhibition in polymeric tubing is primarily due to component adsorption to surfaces.
  • Inhibition increases with tubing length and reduced sample volume, not contact time or flow rate.
  • Chemical compatibility of polymers with DNA dyes is critical for microfluidic device development.