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

Related Concept Videos

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...

You might also read

Related Articles

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

Sort by

Same author

Severe anaemia and invasive bacterial infections in Kenyan children: a 26-year hospital surveillance observational study.

The Lancet. Global health·2026

Same author

Expecting the unexpected: the next outbreak will not necessarily look like the last.

The New Zealand medical journal·2026

Same author

Serious Bacterial Infections in Hospitalized Neonates in Eastern Ethiopia: Investigating the Emerging Pathogen Pantoea dispersa Compared With Klebsiella pneumoniae.

Tropical medicine & international health : TM & IH·2026

Same author

Maternal infection at delivery and risk of stillbirth and early neonatal death: nested case-control studies in East African pregnancy cohorts.

BMC pregnancy and childbirth·2026

Same author

Deaths with preceding hospitalisations within 180 days in eight countries in sub-Saharan Africa and South Asia: A secondary descriptive analysis of the Child Health and Mortality Prevention Surveillance (CHAMPS) network.

BMJ open·2026

Same author

Age-Specific Concentrations and Seroprevalence of Antibodies Against Salmonella Enteritidis (O:9) and Salmonella Typhimurium (O:4,5) Across 3 Sites in Kenya.

The Journal of infectious diseases·2026

Same journal

Identification of eyeworm (<i>Oxyspirura petrowi</i>) and caecal worm (<i>Aulonocephalus pennula</i>) infection levels in Northern bobwhite quail (<i>Colinus virginianus</i>) of the Rolling Plains, TX using a mobile research laboratory: Implications for regional surveillance.

Biomolecular detection and quantification·2019

Same journal

For what factors should we normalize urinary extracellular mRNA biomarkers?

Biomolecular detection and quantification·2019

Same journal

The kinetic requirements of extreme qPCR.

Biomolecular detection and quantification·2019

Same journal

Corrigendum to "Considerations and quality controls when analyzing cell-free tumor DNA" [Biomol. Detect. Quantif. 17 (2019) 100078].

Biomolecular detection and quantification·2019

Same journal

Comparing small urinary extracellular vesicle purification methods with a view to RNA sequencing-Enabling robust and non-invasive biomarker research.

Biomolecular detection and quantification·2019

Same journal

qPCR data analysis: Better results through iconoclasm.

Biomolecular detection and quantification·2019

See all related articles

Search research articles

Related Experiment Video

Updated: Mar 11, 2026

Investigating the Effects of Probiotics on Pneumococcal Colonization Using an In Vitro Adherence Assay

Investigating the Effects of Probiotics on Pneumococcal Colonization Using an In Vitro Adherence Assay

Published on: April 28, 2014

Making standards for quantitative real-time pneumococcal PCR.

Susan C Morpeth¹, Jim F Huggett², David R Murdoch³

¹KEMRI-Wellcome Trust Research Programme, Kilifi, Kenya; Nuffield Department of Clinical Medicine, University of Oxford, Oxford, United Kingdom.

Biomolecular Detection and Quantification

|November 30, 2016

Summary

This summary is machine-generated.

Quantitative PCR (polymerase chain reaction) using the lytA gene for Streptococcus pneumoniae detection showed discrepancies between colony-forming units and genome-copies. This highlights challenges in achieving accurate absolute quantification in diagnostic assays.

Keywords:

Quantitative Real-time PCR Standards Streptococcus pneumoniae

More Related Videos

Characterization of Inflammatory Responses During Intranasal Colonization with Streptococcus pneumoniae

Characterization of Inflammatory Responses During Intranasal Colonization with Streptococcus pneumoniae

Published on: January 17, 2014

High-throughput Quantitative Real-time RT-PCR Assay for Determining Expression Profiles of Types I and III Interferon Subtypes

High-throughput Quantitative Real-time RT-PCR Assay for Determining Expression Profiles of Types I and III Interferon Subtypes

Published on: March 24, 2015

Related Experiment Videos

Last Updated: Mar 11, 2026

Investigating the Effects of Probiotics on Pneumococcal Colonization Using an In Vitro Adherence Assay

Investigating the Effects of Probiotics on Pneumococcal Colonization Using an In Vitro Adherence Assay

Published on: April 28, 2014

Characterization of Inflammatory Responses During Intranasal Colonization with Streptococcus pneumoniae

Characterization of Inflammatory Responses During Intranasal Colonization with Streptococcus pneumoniae

Published on: January 17, 2014

High-throughput Quantitative Real-time RT-PCR Assay for Determining Expression Profiles of Types I and III Interferon Subtypes

High-throughput Quantitative Real-time RT-PCR Assay for Determining Expression Profiles of Types I and III Interferon Subtypes

Published on: March 24, 2015

Area of Science:

Microbiology
Molecular Biology
Infectious Diseases

Background:

Quantitative lytA PCR is crucial for detecting Streptococcus pneumoniae.
In-house standards are commonly used for lytA PCR assays.
Variability in quantification methods can impact diagnostic accuracy.

Purpose of the Study:

To investigate the equivalence of quantifying Streptococcus pneumoniae using colony-forming units (CFU) versus genome-copies via quantitative lytA PCR.
To assess the variability and potential discrepancies between these two quantification methods.

Main Methods:

A standard suspension of Streptococcus pneumoniae was prepared.
Quantification was performed using both colony-forming unit (CFU) counts and genome-copy (GC) measurements.
Quantitative lytA PCR was employed for genome-copy determination.

Main Results:

The median ratio of CFU to genome-copies was 0.19 (IQR: 0.1-1.2).
Genome-copy measurements exhibited lower variability compared to CFU counts.
A significant discrepancy was observed between the CFU and genome-copy quantification methods.

Conclusions:

Equivalence between CFU and genome-copy quantification for Streptococcus pneumoniae using lytA PCR was not consistently demonstrated.
The observed discrepancy highlights inherent challenges in the absolute quantification of bacterial loads.
Further optimization of standardization is needed for reliable quantitative PCR diagnostic assays.