1. Identification of housekeeping genes for microRNA expression analysis in kidney tissues of Pkd1 deficient mouse models.
- Author
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Muñoz JJ, Anauate AC, Amaral AG, Ferreira FM, Meca R, Ormanji MS, Boim MA, Onuchic LF, and Heilberg IP
- Subjects
- Animals, Haploinsufficiency, Mice, Protein Kinase C genetics, Gene Expression Profiling, Genes, Essential genetics, Kidney metabolism, MicroRNAs genetics, Protein Kinase C deficiency
- Abstract
Polycystic kidney disease is a complex clinical entity which comprises a group of genetic diseases that leads to renal cyst development. We evaluated the most suitable housekeeping genes for microRNA expression by RT-qPCR analyses of kidney tissues in Pkd1-deficient mouse models from a panel of five candidates genes (miR-20a, miR-25, miR-26a, miR-191 and U6) and 3 target genes (miR-17, miR-21 and let-7a) using samples from kidneys of cystic mice (Pkd1
flox/flox :Nestincre , CY), non-cystic controls (Pkd1flox/flox , NC), Pkd1-haploinsufficient (Pkd1+/- , HT), wild-type controls (Pkd1+/+ , WT), severely cystic mice (Pkd1V/V , SC), wild-type controls (CO). The stability of the candidate genes was investigated using NormFinder, GeNorm, BestKeeper, DataAssist, and RefFinder software packages and the comparative ΔCt method. The analyses identified miR-26a as the most stable housekeeping gene for all kidney samples, miR-20a for CY and NC, miR-20a and miR-26a for HT and WT, and miR-25 and miR-26a for SC and CO. Expression of miR-21 was upregulated in SC compared to CO and trends of miR-21 upregulation and let-7a downregulation in CY and HT compared to its control kidneys, when normalized by different combinations of miR-20a, miR-25 and miR-26a. Our findings established miR-20a, miR-25, and miR-26a as the best housekeeping genes for miRNA expression analyses by RT-qPCR in kidney tissues of Pkd1-deficient mouse models.- Published
- 2020
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