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Identification of reference micro RNA s for quantitative expression analysis in porcine peripheral blood mononuclear cells treated with polyinosinic–polycytidylic acid
Author(s) -
Wang H.,
Wang J.,
Sun S.,
Wang Y.,
Guo J.,
Ning C.,
Yang K.,
Liu J. F.
Publication year - 2015
Publication title -
international journal of immunogenetics
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.41
H-Index - 47
eISSN - 1744-313X
pISSN - 1744-3121
DOI - 10.1111/iji.12198
Subject(s) - peripheral blood mononuclear cell , reference genes , rna , real time polymerase chain reaction , gene expression , microbiology and biotechnology , microrna , messenger rna , biomarker , gene , biology , in vitro , biochemistry
Summary Peripheral blood mononuclear cells ( PBMC s) are clinically important cells. Detection of micro RNA s (mi RNA s) expression in PBMC s can be useful for mi RNA biomarker discovery for various diseases. Quantitative real‐time PCR ( qRT ‐ PCR ) has become an important method used for measuring mi RNA s expression. However, the reliability of qRT ‐ PCR data critically depends on proper selection of reference genes. Here, we performed qRT ‐ PCR to quantify the expression levels of nine mi RNA s (Ssc‐miR‐16, Hsa‐miR‐25, Ssc‐miR‐34a, Hsa‐miR‐93, Bta‐miR‐92b, Ssc‐miR‐103, Ssc‐miR‐106a, Ssc‐miR‐128 and Ssc‐miR‐107) and one small nuclear RNA (U6) in PBMC s treated with polyinosinic–polycytidylic acid [poly (I:C)] that widely used for simulating viral infection. We used the four statistical algorithms (GeNorm 3.5, NormFinder, BestKeeper and comparative ∆ Ct method) to evaluate gene expression stability and observed that Ssc‐miR‐34a was the best single reference gene and the pair of Ssc‐miR‐107 and Ssc‐miR‐103 was the best combination of reference mi RNA s for porcine PBMC s treated with poly (I:C). Our study shows the first evidence of careful selection of reference mi RNA s in porcine PBMC s and maybe helpful for discovering mi RNA biomarkers for double‐stranded RNA ‐induced disease.