Zhang Ying, Zeng Luying, Wei Yongji, Zhang Ming, Pan Weidong, Sword Gregory A, Yang Fei, Chen Fajun, Wan Guijun
Department of Entomology, College of Plant Protection, Nanjing Agricultural University, Nanjing, China.
Key Laboratory of Plant Health & Crop Safety, Nanjing Agricultural University, Nanjing, China.
Front Physiol. 2022 Aug 8;13:954228. doi: 10.3389/fphys.2022.954228. eCollection 2022.
Manipulating the hypomagnetic field (HMF), which is the absence or significant weakening (<5 μT) of the geomagnetic field (GMF), offers a unique tool to investigate magnetic field effects on organismal physiology, development, behavior and life history. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) has been utilized to study changes in gene expression associated with exposure to the HMF. However, selecting appropriate reference genes (RGs) with confirmed stable expression across environments for RT-qPCR is often underappreciated. Using three algorithms (BestKeeper, NormFinder, and GeNorm), we investigated the expression stability of eight candidate RGs when exposed to the HMF condition versus local GMF during developmental from juveniles to adults in the migratory insect pest, the brown planthopper . During the nymphal stage, & & , & , & , and & were suggested as the most stable RG sets in the 1st to 5th instars, respectively. For 1- to 3-day-old adults, & , & & and & , & , were the optimal RG sets for macropterous and brachypterous females, respectively. & & & and & & & were the optimal RG sets for macropterous and brachypterous males, respectively. These results will facilitate accurate gene expression analyses under the HMF in . The verification approach illustrated in this study highlights the importance of identifying reliable RGs for future empirical studies of magnetobiology (including magnetoreception) that involve magnetic field intensity as a factor.
操控低磁场（HMF），即地磁场（GMF）缺失或显著减弱（<5 μT），为研究磁场对生物体生理、发育、行为和生活史的影响提供了一种独特工具。逆转录定量聚合酶链反应（RT-qPCR）已被用于研究与暴露于HMF相关的基因表达变化。然而，选择在不同环境中具有稳定表达的合适内参基因（RGs）用于RT-qPCR往往未得到充分重视。我们使用三种算法（BestKeeper、NormFinder和GeNorm），研究了在迁飞性害虫褐飞虱从若虫发育到成虫的过程中，暴露于HMF条件与当地GMF条件下时，八个候选内参基因的表达稳定性。在若虫阶段，分别建议在1至5龄若虫中，&&、& &、& &、& &和& &作为最稳定的内参基因组合。对于1至3日龄的成虫，& &、& & &和& &、& &、分别是长翅型和短翅型雌虫的最佳内参基因组合。& & & &和& & & &分别是长翅型和短翅型雄虫的最佳内参基因组合。这些结果将有助于在褐飞虱的HMF条件下进行准确的基因表达分析。本研究中所示的验证方法突出了识别可靠内参基因对于未来涉及磁场强度作为一个因素的磁生物学（包括磁感受）实证研究的重要性。
