Department of Entomology, China Agricultural University, Beijing 100094, China.
Department of Entomology, Henan Institute of Science and Technology, Xinxiang 453003, China.
J Insect Sci. 2017 Jan 1;17(2). doi: 10.1093/jisesa/iex007.
More and more studies have focused on microRNAs (miRNAs) expression in the pest Helicoverpa armigera (Lepidoptera: Noctuidae) recently. Quantitative real-time PCR (qRT-PCR) is being widely used in miRNA expression studies. Suitable reference genes are necessary for the correct analysis of results. In this study, 10 candidate genes of H. armigera were selected and analyzed for their expression stability under different biotic and abiotic conditions with 3 statistical methods, including geNorm, NormFinder, and Bestkeeper. Combination the best number of reference genes was calculated by geNorm. One target gene, let-7, was used to validate the selection of reference genes. The suitable candidate reference genes were shown as follows: miR-9 and U6 snRNA for developmental stages, miR-100 and U6 snRNA for larval tissues, miR-100 and miR-305 for adult tissues, miR-9 and miR-279 for parasitic treatment, miR-998 and U6 snRNA for nuclear polyhedrosis virus infection, miR-9 and U6 snRNA for insecticide treatment, miR-92a, miR-100, and miR-279 for temperature treatment, miR-92a, miR-305, and miR-998 for starvation treatment, miR-9 and miR-279 for light treatment, miR-305 and miR-998 for hormone treatment, and there was not one reference gene suitable for all samples. This study could promote future research on miRNAs expression in H. armigera with optimal reference genes under different experimental conditions.
最近，越来越多的研究集中在鳞翅目夜蛾科害虫棉铃虫（Helicoverpa armigera）的 microRNAs（miRNAs）表达上。定量实时 PCR（qRT-PCR）广泛应用于 miRNA 表达研究。合适的参考基因对于正确分析结果是必要的。在这项研究中，选择了 10 个棉铃虫候选基因，并使用 3 种统计方法（geNorm、NormFinder 和 BestKeeper）分析了它们在不同生物和非生物条件下的表达稳定性。geNorm 计算了最佳参考基因数量的组合。用 let-7 验证了参考基因的选择。合适的候选参考基因如下：miR-9 和 U6 snRNA 用于发育阶段，miR-100 和 U6 snRNA 用于幼虫组织，miR-100 和 miR-305 用于成虫组织，miR-9 和 miR-279 用于寄生处理，miR-998 和 U6 snRNA 用于核多角体病毒感染，miR-9 和 U6 snRNA 用于杀虫剂处理，miR-92a、miR-100 和 miR-279 用于温度处理，miR-92a、miR-305 和 miR-998 用于饥饿处理，miR-9 和 miR-279 用于光照处理，miR-305 和 miR-998 用于激素处理，没有一个参考基因适合所有样本。本研究为在不同实验条件下使用最佳参考基因促进棉铃虫 miRNA 表达的未来研究提供了参考。
