Identification of Suitable Reference Genes for qRT-PCR Analysis of Circulating microRNAs in Hepatitis B Virus-Infected Patients
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Circulating microRNAs (miRNAs) were found to exist in serum/plasma in a highly stable, cell-free form, and aberrantly expressed in many human diseases. Currently, the expression levels of circulating miRNAs are estimated by quantitative real-time polymerase chain reaction. However, no study has systematically evaluated reference genes for evaluating circulating microRNA expression. This study describes the identification and characterization of an appropriate reference gene for the normalization of circulating miRNA levels in hepatitis B virus (HBV)-infected patients and healthy people. Ten miRNAs that resemble the mean expression of the TaqMan low density array together with U6, RNU6B, and miR-16 were validated with two algorithms, geNorm, and NormFinder, after ensuring their equivalent expression between the two study groups. The combination of miR-26a, miR-221, and miR-22* is recommended as the most stable set of reference genes for circulating miRNA evaluation in HBV patients and healthy people.
KeywordsHepatitis B virus Circulating microRNA Quantitative real-time PCR Reference genes Normalization
Hepatitis B virus
Hepatitis C virus
Quantitative reverse-transcription polymerase chain reaction
This article was supported by grants from the National Key Projects for Infectious Disease of China (2008ZX10002-021), the Program of Shanghai Chief Scientist (08XD1400800), the National Natural Science Foundation of China (30700991), and the Research Fund for New Teachers of Fudan University.
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