Abstract
We assessed the expression stability of several messenger (m)RNAs and micro (mi)RNAs from liver and head kidney of rainbow trout Oncorhynchus mykiss using high-throughput RNA sequencing (RNA-seq) and miRNA-seq data. Additionally, four commonly used reference genes and one small non-coding RNA (u6) were also selected to identify ideal reference mRNAs and miRNAs for quantitative real-time (qrt)-PCR analysis of heat stress responses. GeNorm, NormFinder, BestKeeper and comparative ΔCt were employed for analysis of qrt-PCR data to systematically assess the expression stability of candidate mRNAs and miRNAs and stability was ranked using geometric means. β-actin and ef1-α were the most stably expressed reference mRNAs in liver and head kidney, respectively and ssa-mir-26a-5p and ssa-mir-462b-5p were the most stably expressed miRNAs in these tissues. This is the first identification of appropriate reference mRNAs and miRNAs for qrt-PCR analysis of O. mykiss under heat stress.
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