Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Search research articles

Related Experiment Video

Updated: May 31, 2026

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

Stem-loop RT-qPCR for miRNAs.

Martha F Kramer¹

¹Harvard Medical School, Boston, Massachusetts, USA.

Current Protocols in Molecular Biology

|July 7, 2011

Summary

This summary is machine-generated.

This study introduces a novel quantitative reverse-transcription PCR (RT-qPCR) method optimized for accurately measuring short microRNAs (miRNAs) in biological samples. The assay enhances specificity and sensitivity for reliable miRNA quantification.

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by

Same author

Herpes simplex virus 1 microRNAs expressed abundantly during latent infection are not essential for latency in mouse trigeminal ganglia.

Virology·2011

Same author

Numerous conserved and divergent microRNAs expressed by herpes simplex viruses 1 and 2.

Journal of virology·2010

Same author

Enzymatic amplification of DNA by PCR: standard procedures and optimization.

Current protocols in cytometry·2008

Same author

MicroRNAs expressed by herpes simplex virus 1 during latent infection regulate viral mRNAs.

Nature·2008

Same author

Enzymatic amplification of DNA by PCR: standard procedures and optimization.

Current protocols in immunology·2008

Same author

The polymerase chain reaction.

Current protocols in protein science·2008

Same journal

Nondenaturing Polyacrylamide Gel Electrophoresis: Preparation and Analysis of DNA.

Current protocols in molecular biology·2021

Same journal

Purification and Concentration of DNA from Aqueous Solutions: Preparation and Analysis of DNA.

Current protocols in molecular biology·2021

Same journal

Expression of Proteins Using Semliki Forest Virus Vectors: Protein Expression.

Current protocols in molecular biology·2021

Same journal

Methylation and Uracil Interference Assays for Analysis of Protein-DNA Interactions: DNA-Protein Interactions.

Current protocols in molecular biology·2021

Same journal

Separation of Double- and Single-Stranded Nucleic Acids Using Hydroxylapatite Chromatography: Preparation and Analysis of DNA.

Current protocols in molecular biology·2021

Same journal

Pulsed-Field Gel Electrophoresis: Preparation and Analysis of DNA.

Current protocols in molecular biology·2021

See all related articles

Area of Science:

Molecular Biology
Genetics
Biochemistry

Background:

MicroRNAs (miRNAs) are short non-coding RNA molecules crucial for gene regulation.
Standard quantitative PCR (qPCR) methods are unsuitable for miRNA detection due to their small size (17-24 nucleotides).

Purpose of the Study:

To develop a specific and sensitive quantitative reverse-transcription PCR (RT-qPCR) assay for individual miRNA measurement.
To overcome the length limitations of standard PCR for miRNA analysis.

Main Methods:

Utilized a modified RT-qPCR approach with specially designed nucleic acid reagents.
Incorporated a stem-loop RT-primer to lengthen the target cDNA.
Optimized forward and reverse PCR primers and a hydrolysis probe with a minor groove binding (MGB) moiety for enhanced specificity and sensitivity.

More Related Videos

Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells

Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells

Published on: February 21, 2014

High Throughput MicroRNA Profiling: Optimized Multiplex qRT-PCR at Nanoliter Scale on the Fluidigm Dynamic ArrayTM IFCs

High Throughput MicroRNA Profiling: Optimized Multiplex qRT-PCR at Nanoliter Scale on the Fluidigm Dynamic ArrayTM IFCs

Published on: August 3, 2011

Related Experiment Videos

Last Updated: May 31, 2026

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

Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells

Profiling of Estrogen-regulated MicroRNAs in Breast Cancer Cells

Published on: February 21, 2014

High Throughput MicroRNA Profiling: Optimized Multiplex qRT-PCR at Nanoliter Scale on the Fluidigm Dynamic ArrayTM IFCs

High Throughput MicroRNA Profiling: Optimized Multiplex qRT-PCR at Nanoliter Scale on the Fluidigm Dynamic ArrayTM IFCs

Published on: August 3, 2011

Main Results:

The developed assay successfully measures individual miRNAs with high specificity and sensitivity.
The method addresses the challenge of amplifying very short nucleic acid targets.

Conclusions:

This optimized RT-qPCR assay provides a reliable tool for accurate miRNA quantification in various biological samples.
The assay design overcomes previous limitations in detecting short RNA molecules.