title: Reference genes for the relative quantification of microRNAs in renal cell carcinomas and their metastases.
abstract: To obtain accurate results in miRNA expression changes between different sample sets using real-time quantitative polymerase chain reaction (RT-qPCR) analyses, normalization to reference genes that are stably expressed across the sample sets is generally used. A literature search of miRNA expression studies in renal cell carcinoma (RCC) proved that non-miRNAs such as small RNAs or mRNAs have most frequently been used without preceding validation of their suitability. In this study, the most stably expressed miRNAs were ascertained from microarray-based data of miRNA expression in nonmalignant and malignant samples from clear cell RCC and from corresponding distant RCC metastases using the geNorm and NormFinder algorithms. Validation experiments with RT-qPCR were performed for the four best-ranked miRNAs (miR-28, miR-103, miR-106a, miR-151) together with the small RNU6B, RNU44, and RNU48 mostly described in literature. miR-28, miR-103, miR-106a, and RNU48 were proved as the most stably expressed genes. miR-28 is recommended as normalizer if only a single reference gene can be used, while the combinations of miR-28 and miR-103 or of miR-28, miR-103, and miR-106a, respectively, are preferred. RNU6B most frequently used as normalizer in miRNA expression studies should be abandoned in order to avoid misleading results.
authors: Department of Biology, Chemistry and Pharmacy, Free University, Berlin, Germany.
authors: Zofia
authors: Z
authors: Wotschofsky
authors: Helmuth-Alexander
authors: HA
authors: Meyer
authors: Monika
authors: M
authors: Jung
authors: Annika
authors: A
authors: Fendler
authors: Ina
authors: I
authors: Wagner
authors: Carsten
authors: C
authors: Stephan
authors: Jonas
authors: J
authors: Busch
authors: Andreas
authors: A
authors: Erbersdobler
authors: Alexander C
authors: AC
authors: Disch
authors: Hans-Joachim
authors: HJ
authors: Mollenkopf
authors: Klaus
authors: K
authors: Jung
mesh_terms: D002292:Carcinoma, Renal Cell / Q000235:genetics* / Q000556:secondary; D015972:Gene Expression Regulation, Neoplastic; D052138:Genes, Neoplasm; D006801:Humans; D007680:Kidney Neoplasms / Q000235:genetics* / Q000473:pathology; D035683:MicroRNAs / Q000032:analysis* / Q000235:genetics; D009362:Neoplasm Metastasis; D020411:Oligonucleotide Array Sequence Analysis; D012342:RNA, Small Nuclear / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction; D012015:Reference Standards
chemical_list: C544749:MIRN103 microRNA, human; C542541:MIRN106 microRNA, human; D035683:MicroRNAs; D012342:RNA, Small Nuclear; C405486:U6 small nuclear RNA