Yang Anpei, Zhang Hang, Bai Weiwei, Ding Ruifeng, Li Weipeng, Li Guangkuo
Institute of Plant Protection, Xinjiang Uygur Autonomous Region Academy of Agricultural Sciences, Urumqi 830091, China.
Key Laboratory of Integrated Pest Management on Crops in Northwestern Oasis, Ministry of Agriculture and Rural Affairs, National Plant Protection Scientific Observation and Experiment Station of Korla, Urumqi 830091, China.
Insects. 2025 May 15;16(5):527. doi: 10.3390/insects16050527.
The clover cutworm, Rottemberg (Lepidoptera: Noctuidae), is a globally distributed polyphagous pest causing significant economic losses to agricultural crops. RT-qPCR is a gold-standard technique for gene expression analysis, yet its accuracy depends critically on stable reference genes for data normalization. To address the lack of validated reference genes in , we evaluated six candidate genes (, , , , , and ) across four developmental stages (egg, larva, pupa, and adult) and six adult tissues (head, thorax, abdomen, wings, legs, and antennae) using geNorm, NormFinder, BestKeeper, and RefFinder algorithms. Stability analysis identified , , and as the most reliable reference genes for developmental stage normalization, while , , and were validated for adult tissues. Functional validation using the odorant receptor gene revealed significant discrepancies in relative expression levels when normalized with unstable reference genes ( and ), emphasizing the necessity of rigorous reference gene selection. This study establishes the first comprehensive reference gene panel for , providing a robust foundation for gene expression studies in this agriculturally important pest.
苜蓿夜蛾，罗滕贝格（鳞翅目：夜蛾科），是一种全球分布的多食性害虫，给农作物造成重大经济损失。RT-qPCR是基因表达分析的金标准技术，但其准确性关键取决于用于数据标准化的稳定参考基因。为了解决[物种名称未给出]中缺乏经过验证的参考基因的问题，我们使用geNorm、NormFinder、BestKeeper和RefFinder算法，在四个发育阶段（卵、幼虫、蛹和成虫）以及六种成虫组织（头部、胸部、腹部、翅膀、腿部和触角）中评估了六个候选基因（[基因名称未给出]）。稳定性分析确定[基因名称未给出]、[基因名称未给出]和[基因名称未给出]是发育阶段标准化最可靠的参考基因，而[基因名称未给出]、[基因名称未给出]和[基因名称未给出]在成虫组织中得到验证。使用气味受体基因[基因名称未给出]进行的功能验证表明，当用不稳定的参考基因（[基因名称未给出]和[基因名称未给出]）进行标准化时，相对表达水平存在显著差异，强调了严格选择参考基因的必要性。本研究建立了首个针对[物种名称未给出]的全面参考基因库，为这种农业重要害虫的基因表达研究提供了坚实基础。
