Molecular quantification of canine specific microRNA species

MicroRNAs (miRNAs) belong to the heterogeneous group of non-coding, regulatory RNA molecules. They play an important role in physiological functions, especially during tissue development. Moreover, their cell-type specific expression levels are also thought to be altered in association with different pathological conditions, including metabolic disorders and cancer. However, the vast majority of molecular assays for their quantification have been established for humans and mice, with no reports on specific research tools for dogs. Here canine-specific SYBRGreen based quantitative, real-time PCR assays are introduced for the quantification of 21 miRNAs and five small nucleolar RNA (snoRNA). The assay includes a timesaving and economical universal one-tube cDNA synthesis approach which allows for a more robust relative quantification than the commonly used multiple specific reverse transcriptions. GeNorm and Norm-Finder analysis were used for the evaluation of potential housekeeping genes and identified miRNAs to be more stable housekeeping genes by trend. (c) 2013 Elsevier Ltd. All rights reserved.