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Related Concept Videos

Real Time RT-PCR02:57

Real Time RT-PCR

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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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Transcription01:10

Transcription

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Overview
Transcription is the process of synthesizing RNA from a DNA sequence by RNA polymerase. It is the first step in producing a protein from a gene sequence. Additionally, many other proteins and regulatory sequences are involved in the proper synthesis of messenger RNA (mRNA). Regulation of transcription is responsible for the differentiation of all the different types of cells and often for the proper cellular response to environmental signals.
Transcription Can Produce Different Kinds...
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PCR01:32

PCR

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Measurement: Standard Units03:38

Measurement: Standard Units

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Every measurement provides three kinds of information: the size or magnitude of the measurement (a number), a standard of comparison for the measurement (a unit), and an indication of the uncertainty of the measurement. While the number and unit are explicitly represented when a quantity is written, the uncertainty is an aspect of the errors in the measurement results.
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Transcription Factors02:16

Transcription Factors

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Tissue-specific transcription factors contribute to diverse cellular functions in mammals. For example, the gene for beta globin, a major component of hemoglobin, is present in all cells of the body. However, it is only expressed in red blood cells because the transcription factors that can bind to the promoter sequences of the beta globin gene are only expressed in these cells. Tissue-specific transcription factors also ensure that mutations in these factors may impair only the function of...
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Master Transcription Regulators02:23

Master Transcription Regulators

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Master transcription regulators are regulatory proteins that are predominantly responsible for regulating the expression of multiple genes. Often these genes work in concert to drive a  complex process. Activation of a master transcription regulator can lead to a cascade of transcriptional activation necessary for that outcome. These regulators can directly bind to the regulatory sequences of the various genes involved, or they can indirectly regulate transcription by binding to regulatory...
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Quantitative Real-Time PCR using the Thermo Scientific Solaris qPCR Assay
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Real-Time Reverse Transcription Quantitative PCR (RT-qPCR) Methodological Standards and Reporting Practices.

Stephen A Bustin1, Carl T Wittwer2

  • 1Medical Technology Research Centre, Faculty of Health, Education, Medicine and Social Care, Anglia Ruskin University, Chelmsford, United Kingdom.

Clinical Chemistry
|January 28, 2026
PubMed
Summary
This summary is machine-generated.

Methodological reporting in real-time reverse transcription quantitative PCR (RT-qPCR) assays remains poor, with critical parameters like RNA integrity and PCR efficiency declining significantly. Adherence to Minimum Information for Publication of Quantitative PCR Experiments (MIQE) guidelines is insufficient, impacting assay reproducibility.

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Area of Science:

  • Life Sciences
  • Molecular Biology
  • Biotechnology

Background:

  • Concerns persist regarding the methodological quality and reporting transparency of real-time reverse transcription quantitative PCR (RT-qPCR).
  • The COVID-19 pandemic highlighted these issues in diagnostic testing.
  • The Minimum Information for Publication of Quantitative PCR Experiments (MIQE) guidelines, updated in 2025, have had a modest impact on standardizing assay reporting.

Purpose of the Study:

  • To assess trends in RT-qPCR methodological reporting between 2007 and 2025.
  • To evaluate the impact of MIQE guidelines on reporting quality.
  • To compare reporting practices across different time points, regions, and MIQE citation status.

Main Methods:

  • PubMed Central searches and manual evaluation of 355 full-text articles from 2019 and 2024.
  • Analysis of parameters including RNA integrity, oligonucleotide disclosure, reference gene validation, and PCR efficiency.
  • Evaluation of targeted cohorts focusing on reference genes and PCR efficiency.

Main Results:

  • Reporting of core RT-qPCR parameters remained low or declined between 2019 and 2024.
  • RNA integrity reporting decreased from 22% to 11%, reference gene validation from 13% to 5%, and PCR efficiency reporting from 13% to 1%.
  • MIQE-citing papers in 2024 showed better adherence but still omitted essential details; Asia dominates RT-qPCR output, while Europe leads in MIQE citations.

Conclusions:

  • Incomplete experimental design and reporting in RT-qPCR assays continue to compromise reproducibility and robustness.
  • Despite updated MIQE guidelines, adherence to essential reporting standards remains inadequate.
  • Further efforts are needed to improve methodological quality and transparency in RT-qPCR publications.