Identification and evaluation of reference genes for qRT-PCR studies in <i>Lentinula edodes</i>

<div><p><i>Lentinula edodes</i> (shiitake mushroom) is a common edible mushroom with a number of potential therapeutic and nutritional applications. It contains various medically important molecules, such as polysaccharides, terpenoids, sterols, and lipids, were contained in this mushroom. Quantitative real-time polymerase chain reaction (qRT-PCR) is a powerful tool to analyze the mechanisms underlying the biosynthetic pathways of these substances. qRT-PCR is used for accurate analyses of transcript levels owing to its rapidity, sensitivity, and reliability. However, its accuracy and reliability for the quantification of transcripts rely on the expression stability of the reference genes used for data normalization. To ensure the reliability of gene expression analyses using qRT-PCR in <i>L</i>. <i>edodes</i> molecular biology research, it is necessary to systematically evaluate reference genes. In the current study, ten potential reference genes were selected from <i>L</i>. <i>edodes</i> genomic data and their expression levels were measured by qRT-PCR using various samples. The expression stability of each candidate gene was analyzed by three commonly used software packages: geNorm, NormFinder, and BestKeeper. Base on the results, <i>Rpl4</i> was the most stable reference gene across all experimental conditions, and <i>Atu</i> was the most stable gene among strains. <i>18S</i> was found to be the best reference gene for different development stages, and <i>Rpl4</i> was the most stably expressed gene under various nutrient conditions. The present work will contribute to qRT-PCR studies in <i>L</i>. <i>edodes</i>.</p>