A novel qPCR assay for viral encoded microRNAs

Irit Reichenstein; Natalie Aizenberg; Maya Goshen; Zvi Bentwich; Yonat Shemer Avni

doi:10.1016/j.jviromet.2009.10.018

A novel qPCR assay for viral encoded microRNAs

Irit Reichenstein, Natalie Aizenberg, Maya Goshen, Zvi Bentwich, Yonat Shemer Avni^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

Abstract

MicroRNAs (miRNAs) are short non-coding RNAs that postranscriptionally regulate viral and host gene expression. Reliable and simple assays for detecting and analyzing miRNAs during viral infections are critical for clinical and research purposes. A highly sensitive quantitative real-time PCR (qPCR) assay was developed. This approach, using a generic hydrolysis probe, detects and quantifies miRNAs in cell culture and in clinical samples obtained from patients. The assay is based on preparation of cDNA libraries by polyadenylation of total RNA and reverse transcription, followed by detection of specific miRNAs in the cDNA library by qPCR. The qPCR test was highly sensitive and specific, distinguishing between miRNAs that differ by as little as a single nucleotide with remarkable reproducibility. When applied to clinical samples the assay could detect the differential expression of EBV encoded miRNAs in peripheral blood cells of healthy EBV carriers and patients with acute EBV infection, which makes it a powerful tool for the study of differential expression of miRNAs in health and during viral infections.

Original language	English
Pages (from-to)	323-328
Number of pages	6
Journal	Journal of Virological Methods
Volume	163
Issue number	2
DOIs	https://doi.org/10.1016/j.jviromet.2009.10.018
State	Published - Feb 2010
Externally published	Yes

Keywords

EBV
Hydrolysis probe
MicroRNA
qPCR
Real-time PCR
Virus

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10.1016/j.jviromet.2009.10.018

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title = "A novel qPCR assay for viral encoded microRNAs",

abstract = "MicroRNAs (miRNAs) are short non-coding RNAs that postranscriptionally regulate viral and host gene expression. Reliable and simple assays for detecting and analyzing miRNAs during viral infections are critical for clinical and research purposes. A highly sensitive quantitative real-time PCR (qPCR) assay was developed. This approach, using a generic hydrolysis probe, detects and quantifies miRNAs in cell culture and in clinical samples obtained from patients. The assay is based on preparation of cDNA libraries by polyadenylation of total RNA and reverse transcription, followed by detection of specific miRNAs in the cDNA library by qPCR. The qPCR test was highly sensitive and specific, distinguishing between miRNAs that differ by as little as a single nucleotide with remarkable reproducibility. When applied to clinical samples the assay could detect the differential expression of EBV encoded miRNAs in peripheral blood cells of healthy EBV carriers and patients with acute EBV infection, which makes it a powerful tool for the study of differential expression of miRNAs in health and during viral infections.",

keywords = "EBV, Hydrolysis probe, MicroRNA, qPCR, Real-time PCR, Virus",

author = "Irit Reichenstein and Natalie Aizenberg and Maya Goshen and Zvi Bentwich and Avni, {Yonat Shemer}",

note = "Funding Information: We wish to thank Dr. S. Gilad and Dr. E. Meiri, for helpful discussions and Prof. Maureen G. Friedman for the enlightening editorial remarks. This work was supported by The Israel Science Foundation [grant no. 1375/05 ]; and The Israel Ministry for Science, Culture, and Sport [grant no. 3-4435 ]. Appendix A ",

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doi = "10.1016/j.jviromet.2009.10.018",

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volume = "163",

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AU - Reichenstein, Irit

AU - Aizenberg, Natalie

AU - Goshen, Maya

AU - Bentwich, Zvi

AU - Avni, Yonat Shemer

N1 - Funding Information: We wish to thank Dr. S. Gilad and Dr. E. Meiri, for helpful discussions and Prof. Maureen G. Friedman for the enlightening editorial remarks. This work was supported by The Israel Science Foundation [grant no. 1375/05 ]; and The Israel Ministry for Science, Culture, and Sport [grant no. 3-4435 ]. Appendix A

PY - 2010/2

Y1 - 2010/2

N2 - MicroRNAs (miRNAs) are short non-coding RNAs that postranscriptionally regulate viral and host gene expression. Reliable and simple assays for detecting and analyzing miRNAs during viral infections are critical for clinical and research purposes. A highly sensitive quantitative real-time PCR (qPCR) assay was developed. This approach, using a generic hydrolysis probe, detects and quantifies miRNAs in cell culture and in clinical samples obtained from patients. The assay is based on preparation of cDNA libraries by polyadenylation of total RNA and reverse transcription, followed by detection of specific miRNAs in the cDNA library by qPCR. The qPCR test was highly sensitive and specific, distinguishing between miRNAs that differ by as little as a single nucleotide with remarkable reproducibility. When applied to clinical samples the assay could detect the differential expression of EBV encoded miRNAs in peripheral blood cells of healthy EBV carriers and patients with acute EBV infection, which makes it a powerful tool for the study of differential expression of miRNAs in health and during viral infections.

AB - MicroRNAs (miRNAs) are short non-coding RNAs that postranscriptionally regulate viral and host gene expression. Reliable and simple assays for detecting and analyzing miRNAs during viral infections are critical for clinical and research purposes. A highly sensitive quantitative real-time PCR (qPCR) assay was developed. This approach, using a generic hydrolysis probe, detects and quantifies miRNAs in cell culture and in clinical samples obtained from patients. The assay is based on preparation of cDNA libraries by polyadenylation of total RNA and reverse transcription, followed by detection of specific miRNAs in the cDNA library by qPCR. The qPCR test was highly sensitive and specific, distinguishing between miRNAs that differ by as little as a single nucleotide with remarkable reproducibility. When applied to clinical samples the assay could detect the differential expression of EBV encoded miRNAs in peripheral blood cells of healthy EBV carriers and patients with acute EBV infection, which makes it a powerful tool for the study of differential expression of miRNAs in health and during viral infections.

KW - EBV

KW - Hydrolysis probe

KW - MicroRNA

KW - qPCR

KW - Real-time PCR

KW - Virus

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A novel qPCR assay for viral encoded microRNAs

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Keywords

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