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Real Time RT-PCR

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...

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Updated: Jul 28, 2025

Circulating MicroRNA Quantification Using DNA-binding Dye Chemistry and Droplet Digital PCR

Circulating MicroRNA Quantification Using DNA-binding Dye Chemistry and Droplet Digital PCR

Published on: June 26, 2016

Reverse transcription-free digital-quantitative-PCR for microRNA analysis.

Hao T Mai¹, Brice C Vanness¹, Thomas H Linz¹

¹Department of Chemistry, Wayne State University, 5101 Cass Ave, Detroit, MI 48202, USA. tlinz@wayne.edu.

|June 2, 2023

Summary

This summary is machine-generated.

This study introduces base-stacking digital-quantitative-PCR (BS-dqPCR), a faster method for measuring microRNAs (miRNAs). BS-dqPCR eliminates time-consuming steps, enabling rapid and sensitive quantification of miRNAs for research.

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Last Updated: Jul 28, 2025

Circulating MicroRNA Quantification Using DNA-binding Dye Chemistry and Droplet Digital PCR

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Published on: June 26, 2016

Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs

Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs

Published on: April 14, 2015

Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR

Cerebrospinal Fluid MicroRNA Profiling Using Quantitative Real Time PCR

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Area of Science:

Molecular Biology
Biotechnology
Biomedical Research

Background:

MicroRNAs (miRNAs) are crucial regulators of biological processes, making them key targets in biomedical research.
Low miRNA abundance in samples requires highly sensitive detection methods for routine laboratory analysis.
Current PCR-based miRNA quantification methods are hindered by lengthy ligation and reverse transcription steps.

Purpose of the Study:

To develop a rapid and sensitive method for quantifying microRNAs (miRNAs) in digital microwell arrays.
To overcome the limitations of conventional PCR techniques by eliminating preparatory steps.
To establish a cost-effective and time-efficient approach for routine miRNA measurements.

Main Methods:

Development of base-stacking digital-quantitative-PCR (BS-dqPCR) to directly quantify miRNAs without ligation or reverse transcription.
Implementation of a novel asymmetric thermocycling program to enhance on-target miRNA signals and minimize non-specific amplification.
Evaluation of BS-dqPCR analytical performance across a range of miRNA concentrations.

Main Results:

BS-dqPCR significantly reduces assay time and cost compared to traditional miRNA PCR methods.
The developed asymmetric thermocycling program effectively distinguishes miRNA signals from background noise.
BS-dqPCR demonstrates a clear digital signal increase with rising miRNA copy numbers.
The quantitative PCR dimension of BS-dqPCR accurately measures miRNA levels within a wide dynamic range.

Conclusions:

BS-dqPCR offers a rapid, sensitive, and cost-effective solution for routine microRNA quantification.
This method streamlines miRNA analysis, making it more accessible for research laboratories.
BS-dqPCR shows significant potential for advancing microRNA-based diagnostics and research.