Tissue Specific Reference Genes for MicroRNA Expression Analysis in a Mouse Model of Peripheral Nerve Injury

MicroRNAs (miRNAs) have emerged as master switch regulators in many biological processes in health and disease, including neuropathy. miRNAs are commonly quantified by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), usually estimated as relative expression through r...

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Bibliographic Details
Published in:Frontiers in molecular neuroscience 2019-11, Vol.12, p.283-283
Main Authors: Kalpachidou, Theodora, Kummer, Kai K, Mitrić, Miodrag, Kress, Michaela
Format: Article
Language:English
Subjects:
Animal models
Biomarkers
Dorsal horn
Dorsal root ganglia
Gene expression
medial prefrontal cortex
Medical research
Methods
MicroRNAs
miRNA
miRNA normalization
ncRNA
Neuroscience
Non-coding RNA
Pain
Peripheral nerves
Peripheral neuropathy
Physiology
Polymerase chain reaction
Prefrontal cortex
Reverse transcription
spared nerve injury
Spinal cord
spinal dorsal horn
Surgery
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Summary:MicroRNAs (miRNAs) have emerged as master switch regulators in many biological processes in health and disease, including neuropathy. miRNAs are commonly quantified by reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR), usually estimated as relative expression through reference genes normalization. Different non-coding RNAs (ncRNAs) are used for miRNA normalization; however, there is no study identifying the optimal reference genes in animal models for peripheral nerve injury. We evaluated the stability of eleven ncRNAs, commonly used for miRNA normalization, in dorsal root ganglia (DRG), dorsal horn of the spinal cord (dhSC), and medial prefrontal cortex (mPFC) in the mouse spared nerve injury (SNI) model. After RT-qPCR, the stability of each ncRNA was determined by using four different methods: BestKeeper, the comparative delta-Cq method, geNorm, and NormFinder. The candidates were rated according to their performance in each method and an overall ranking list was compiled. The most stable ncRNAs were: sno420, sno429, and sno202 in DRG; sno429, sno202, and U6 in dhSC; sno202, sno420, and sno142 in mPFC. We provide the first reference genes' evaluation for miRNA normalization in different neuronal tissues in an animal model of peripheral nerve injury. Our results underline the need for careful selection of reference genes for miRNA normalization in different tissues and experimental conditions. We further anticipate that our findings can be used in a broad range of nerve injury related studies, to ensure validity and promote reproducibility in miRNA quantification.
ISSN:1662-5099
1662-5099
DOI:10.3389/fnmol.2019.00283