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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...
66.8K

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Probe-based Real-time PCR Approaches for Quantitative Measurement of microRNAs
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Normalization of circulating microRNA expression data obtained by quantitative real-time RT-PCR.

Francesco Marabita, Paola de Candia, Anna Torri

    Briefings in Bioinformatics
    |August 5, 2015
    PubMed
    Summary

    Identifying stable reference microRNAs (miRNAs) is crucial for accurate blood biomarker research. This study presents a method using algorithms to find reliable normalizers for quantitative real-time PCR (qPCR) assays, improving data consistency.

    Keywords:
    Normfindercirculating miRNAgeNormnormalizationqPCRreference genes

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    Area of Science:

    • Biomarker discovery
    • Molecular diagnostics
    • Genomics and transcriptomics

    Background:

    • Circulating microRNAs (miRNAs) are promising biomarkers for various diseases.
    • Standardization of quantitative real-time reverse transcription polymerase chain reaction (qPCR) data normalization remains a challenge.

    Purpose of the Study:

    • To develop and validate a robust method for identifying stable reference microRNAs (miRNAs) for normalization.
    • To improve the reliability of high-throughput miRNA analysis in blood samples.

    Main Methods:

    • Utilized a combination of algorithms to analyze high-throughput miRNA expression data.
    • Identified candidate reference miRNAs suitable for normalization in both discovery and validation phases.
    • Applied identified normalizers to reduce non-biological variation in qPCR data.

    Main Results:

    • Successfully identified stable reference miRNAs that effectively normalize miRNA expression data.
    • Demonstrated the utility of these normalizers in reducing non-biological variation.
    • Presented case studies illustrating the application in physiological and pathological contexts.

    Conclusions:

    • The discovery of stable reference miRNAs is essential for appropriate normalization of qPCR assays.
    • This methodology enhances the accuracy and reproducibility of circulating miRNA biomarker research.
    • Stable normalizers improve the clinical relevance of miRNA profiling.