Identification and Evaluation of qRT-PCR Reference Genes in .

Appropriate reference genes must be selected for accurate qRT-PCR data to conduct a thorough gene expression analysis in the sorghum aphid (, Hemiptera, Aphididae). This approach will establish a foundation for gene expression analysis and determines the efficacy of RNA interference in the sorghum aphid. Nine potential reference genes, including , , , , , , , , and , were assessed under various experimental conditions to gauge their suitability based on qRT-PCR Ct values. The stability of these candidate reference genes in diverse experimental setups was analyzed employing several techniques, including the ΔCt comparative method, geNorm, Normfinder, BestKeeper, and RefFinder. The findings revealed that the quantity of ideal reference genes ascertained by the geNorm method for diverse experimental conditions remained consistent. For the developmental stages of the sorghum aphid, and proved to be the most dependable reference genes, whereas and were recommended as the most stable reference genes for different tissues. In experiments involving wing dimorphism, and were identified as the optimal reference gene pair. Under varying temperatures, and were found to be the most dependable gene pair. For studies focusing on insecticide susceptibility, and emerged as the most stable candidate reference genes. Across all experimental conditions, and was the optimal combination of reference genes in the sorghum aphid. This research has pinpointed stable reference genes that can be utilized across various treatments, thereby enhancing gene expression studies and functional genomics research on the sorghum aphid.