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Reference Gene Selection for Quantitative Gene Expression Analysis in Argynnis hyperbius
Hong-Juan Xin1, Chen-Yang Liu1, Feng Yan1
1Henan International Laboratory for Green Pest Control, College of Plant Protection, Henan Agricultural University, Zhengzhou 450046, China.
Insects
|October 28, 2025
Summary
This study identified optimal reference genes for studying the environmental indicator species Argynnis hyperbius. AK, EF1α, ACT, RPL32, and RPL27 showed the most stable expression across various conditions, improving RT-qPCR reliability.
Area of Science:
- * Entomology
- * Molecular Biology
- * Environmental Science
Background:
- * Argynnis hyperbius, an environmental indicator species, faces decline due to climate change and environmental pressures.
- * Reliable gene expression studies using RT-qPCR are crucial for understanding its physiological and adaptive responses.
- * Identifying stable reference genes is fundamental for accurate RT-qPCR analysis in A. hyperbius.
Purpose of the Study:
- * To screen and evaluate the expression stability of 10 commonly used reference genes in Argynnis hyperbius.
- * To determine the most suitable reference genes for normalization under different developmental stages, genders, and temperature treatments.
- * To validate the reliability of selected reference genes by analyzing HSP90 expression.
Main Methods:
- * Selection of 10 candidate reference genes (ACT, α-TUB, AK, GAPDH, EF1α, BTF3, RPS3, RPL10, RPL32, RPL27).
- * Evaluation of gene expression stability using the RefFinder tool across different developmental stages, genders, and temperatures.
- * Validation of reference gene stability by analyzing the relative transcription levels of HSP90.
Main Results:
- * Optimal reference gene pairs were identified for different experimental conditions: AK/EF1α (developmental stages), ACT/RPL32 (sexes), and EF1α/RPL27 (temperatures).
- * EF1α and RPL32 were identified as the most stable reference genes overall.
- * Analysis of HSP90 expression using EF1α/RPL32 revealed significant differences at varying temperatures (4 °C vs. 26 °C/37 °C).
Conclusions:
- * The study provides validated reference gene pairs for accurate RT-qPCR analysis in Argynnis hyperbius.
- * Findings enhance the reliability of gene expression studies in this indicator species, aiding research on its adaptation to environmental changes.
- * This research lays the groundwork for further investigations into the physiological and biochemical mechanisms of A. hyperbius.
