Expression stability of candidate RT‐qPCR housekeeping genes in Spodoptera frugiperda (Lepidoptera: Noctuidae)

Abstract Reverse‐transcription quantitative polymerase chain reaction (RT‐qPCR) is commonly used to quantify gene expression. For normalization, the expression of each gene is compared with a reference “housekeeping” gene that is stably expressed under relevant stress. Unfortunately, there have been no reports on the stability of such reference genes under various treatments of the Spodoptera frugiperda . In this study, we used five tools (RefFinder, GeNorm, NormFinder, BestKeeper, and ΔCt methods) to evaluate the stability of 12 candidate reference genes (RPS18, β‐tubulin, GAPDH, RPS7, RPS15, RPL7, RPL32, Actin‐5C, EF1‐α, EF1‐γ, RPL27, and ACE) in different instars, tissues, and treatments (high and low temperature, UV‐A, and emamectin benzoate). Several ribosomal proteins (RPS7, RPS15, RPL32, RPS18, and RPL7), GAPDH, Actin‐5C, and β‐tubulin, were relatively stable, suggesting that they are ideal housekeeping genes for various treatments. ACE was extremely unstable under various experimental treatments, rendering it unsuitable as an internal reference. This study identified the reference housekeeping genes stably expressed by S. frugiperda under different treatments, thus setting a foundation for further exploration of the physiological and biochemical mechanisms.