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Related Concept Videos

Real Time RT-PCR02:57

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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...
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Circulating MicroRNA Quantification Using DNA-binding Dye Chemistry and Droplet Digital PCR
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Droplet digital PCR enabled by microfluidic impact printing for absolute gene quantification.

Yang Pan1, Tuo Ma1, Qi Meng1

  • 1Department of Precision Machinery & Precision Instrumentation, University of Science & Technology of China, Hefei, Anhui, 230027, China; Key Laboratory of Precision Scientific Instrumentation of Anhui Higher Education Institutes, University of Science and Technology of China, Hefei, Anhui, 230027, China.

Talanta
|February 20, 2020
PubMed
Summary

A novel digital droplet PCR method uses a microfluidic impact printer for precise gene expression analysis. This technology accurately quantifies rare biological variants and gene targets in clinical samples.

Keywords:
Digital PCRGenetic analysesMicrofluidic impact printing

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Area of Science:

  • Molecular Biology
  • Biotechnology
  • Genomics

Background:

  • Digital PCR offers high sensitivity for detecting rare biological variants.
  • Accurate quantification of nucleic acids is crucial for various applications, including gene expression analysis.

Purpose of the Study:

  • To introduce and validate a new digital droplet-based PCR technology utilizing a microfluidic impact printer (MIP).
  • To assess the accuracy and applicability of MIP-enabled digital PCR for gene expression analysis in clinical samples.

Main Methods:

  • Partitioning genetic targets into nanoliter droplets using a microfluidic impact printer and disposable chip.
  • Verifying accuracy by detecting GAPDH gene concentration gradients across four orders of magnitude.
  • Applying the technology to detect p53 gene expression in colon cancer and adjacent nontumorous tissues.

Main Results:

  • The MIP-enabled digital PCR accurately quantified GAPDH gene concentrations from 0.464 to 464 copies/μL.
  • Absolute quantification of nucleic acid copies was achieved for p53 gene expression.
  • Results demonstrated consistency with conventional real-time PCR.

Conclusions:

  • The microfluidic impact printer-enabled digital PCR is a highly sensitive and accurate method for quantitative measurements.
  • This technology shows significant potential for detecting gene expression in clinical settings, particularly for cancer research.