Yang Jing, Zhang Yuying, Zhao Jin, Gao Yue, Liu Zhongfang, Zhang Pengjiu, Fan Jianbin, Zhou Xuguo, Fan Renjun
College of Plant Protection, Shanxi Agricultural University (Institute of Plant Protection, Shanxi Academy of Agricultural Science), Shanxi Key Laboratory of Integrated Pest Management in Agriculture, Taiyuan, China.
Department of Entomology, University of Kentucky, Lexington, KY, United States.
Front Physiol. 2020 Apr 21;11:378. doi: 10.3389/fphys.2020.00378. eCollection 2020.
Hawthorn spider mite, Zacher, is an economically important arthropod pest for fruit trees and woody ornamental plants. Extensive and repetitive use of synthetic acaricides has led to the development of resistance in . To understand the molecular basis of pesticide resistance, and to develop genetic-based control alternatives (e.g., RNAi-based biopesticides), a standardized protocol for real-time quantitative reverse transcription PCR (RT-qPCR) is needed. In the proceeding phase of this research, we screened for the internal references for RT-qPCR analysis from a pool of housekeeping genes under the intrinsic conditions, including developmental stage, sex, and diapause. Here, we continued our efforts to search for the reference genes under an array of extrinsic conditions, including temperature, humidity, photoperiod, host plant, and dietary RNAi. The stability of these candidate reference genes was investigated using geNorm, NormFinder, BestKeeper, and ΔCt method, respectively. Finally, RefFinder, a statistical platform integrating all four algorisms, provided a comprehensive list of genes for each extrinsic condition: (1) , α and were the best candidates for temperature, (2) , , and were the most stable genes for humidity, (3) , , and were the top reference genes for photoperiod, (4) , , and α were recommended for host plants, and (5) , , and were the top choices for dietary RNAi. Overall, , , and were the most commonly selected reference genes in regardless of the experimental conditions, including both intrinsic and extrinsic. Information present here lays the foundation for the genomic and functional genomic research in .
山楂叶螨（Zacher）是一种对果树和木本观赏植物具有重要经济影响的节肢动物害虫。广泛且重复使用合成杀螨剂已导致其产生抗药性。为了解农药抗性的分子基础，并开发基于基因的控制替代方法（如基于RNA干扰的生物农药），需要一种标准化的实时定量逆转录PCR（RT-qPCR）方案。在本研究的前期阶段，我们在包括发育阶段、性别和滞育等内在条件下，从一组管家基因中筛选用于RT-qPCR分析的内参基因。在此，我们继续努力在一系列外在条件下寻找内参基因，包括温度、湿度、光周期、寄主植物和饲料RNA干扰。分别使用geNorm、NormFinder、BestKeeper和ΔCt方法研究这些候选内参基因的稳定性。最后，RefFinder这个整合了所有四种算法的统计平台，针对每种外在条件提供了一份综合基因列表：（1） 、α和 是温度条件下的最佳候选基因，（2） 、 和 是湿度条件下最稳定的基因，（3） 、 和 是光周期条件下的最佳内参基因，（4） 、 和α推荐用于寄主植物，（5） 、 和 是饲料RNA干扰条件下的首选基因。总体而言，无论内在还是外在实验条件如何， 都是山楂叶螨中最常被选择的内参基因。本文提供的信息为山楂叶螨的基因组和功能基因组研究奠定了基础。
