School of Basic Medicine and Biological Sciences, Soochow University, Suzhou, Jiangsu, 215123, China.
Sericulture Institute of Soochow University, Suzhou, Jiangsu, 215123, China.
Bull Entomol Res. 2023 Dec;113(6):845-857. doi: 10.1017/S0007485323000536. Epub 2023 Nov 24.
The silkworm (Lepidoptera: Bombycidae) is a lepidopteran model insect of great economic importance. The parasitoid (Diptera, Tachinidae) is the major pest of and also a promising candidate for biological control. However, the molecular interactions between hosts and dipteran parasitoids have only partially been studied. Gene expression analysis by reverse-transcription quantitative real-time polymerase chain reaction (RT-qPCR) is indispensable to characterise their interactions. Accurate normalisation of RT-qPCR-based gene expression requires the use of reference genes that are constantly expressed irrespective of experimental conditions. In this study, the expression stability of 13 traditionally used reference genes was estimated by five statistical algorithms (ΔCt, geNorm, Normfinder, BestKeeper, and RefFinder) to determine the best reference genes for gene expression studies in different tissues of under . parasitism. Specifically, was the best reference gene in epidermis and testis, while was the most stable gene in prothoracic gland and midgut. , , , , and were the most suitable genes in head, silk gland, fat body, haemolymph, Malpighian tubule and ovary, respectively. Our study offers a set of suitable reference genes for gene expression normalisation in under the parasitic stress of . , which will benefit the in-depth exploration of host-dipteran parasitoid interactions, and also provide insights for further improvements of resistance against parasitoids and biocontrol efficacy of dipteran parasitoids.
家蚕（鳞翅目：天蚕科）是一种具有重要经济价值的鳞翅目模式昆虫。寄生蜂（双翅目，寄蝇科）是家蚕的主要害虫，也是生物防治的有前途的候选者。然而，宿主与双翅目寄生蜂之间的分子相互作用仅部分得到研究。通过反转录定量实时聚合酶链反应（RT-qPCR）进行基因表达分析对于描述它们的相互作用是必不可少的。准确地对基于 RT-qPCR 的基因表达进行归一化需要使用无论实验条件如何都持续表达的参考基因。在这项研究中，通过五种统计算法（ΔCt、geNorm、Normfinder、BestKeeper 和 RefFinder）评估了 13 种传统使用的参考基因的表达稳定性，以确定在 寄生胁迫下研究不同组织中的基因表达的最佳参考基因。 在家蚕的表皮和睾丸中是最佳的参考基因，而 在前胸腺和中肠中是最稳定的基因。 、 、 、 、 和 分别是头部、丝腺、脂肪体、血液、马氏管和卵巢中最适合的基因。我们的研究为寄生胁迫下 的基因表达归一化提供了一组合适的参考基因，这将有助于深入探索宿主 - 双翅目寄生蜂相互作用，并为进一步提高家蚕对寄生蜂的抗性和双翅目寄生蜂的生物防治效果提供见解。
