Quantitative real-time polymerase chain reaction (qRT-PCR) is a powerful tool for evaluating gene expression, but its accuracy is affected by the stability of the reference genes used for normalization. The Minimum Information for Publication of Quantitative Real-time PCR Experiments (MIQE) guidelines indicated that it was important to use multiple stable reference genes as compound reference genes for acquiring optimal experimental results. In this study, the expression levels of eight candidate reference genes (SDHA, TBP, GAPDH, etc.) were detected by qRT-PCR in rat long bones at different developmental stages [gestation day (GD) 20, postnatal week (PW) 6 and PW12] under physiological conditions. Software geNorm, NormFinder, and BestKeeper were used to comprehensively evaluate the stability of the eight reference genes for screening out the most stable compound reference genes in each period. Additionally, the pathological model of prenatal dexamethasone exposure (PDE) was used to verify the stability and reliability of the selected compound reference genes. The result showed that two reference genes as compound reference genes for normalization were optimal. In the intrauterine period, SDHA and TBP could be selected as the compound reference genes, while YWHAZ and GAPDH could be selected at PW6 and PW12, and there was no significant gender difference in the selection of reference genes. The above compound reference genes remained stable in the PDE model and could make the statistical significance of the experimental results more remarkable. In conclusion, this study screened out the optimal compound reference genes in rat long bones before and after birth.

定量实时聚合酶链反应 (qRT-PCR) 是评估基因表达的强大工具，但其准确性受用于标准化的参考基因的稳定性影响。定量实时 PCR 实验 (MIQE) 发布的最低信息指南表明，使用多个稳定的参考基因作为复合参考基因以获得最佳实验结果非常重要。本研究通过qRT-PCR检测了大鼠不同发育阶段长骨中8个候选参考基因（SDHA、TBP、GAPDH等）的表达水平[妊娠天（GD）20，产后周（PW）6和 PW12] 在生理条件下。软件 geNorm、NormFinder、和BestKeeper综合评价8个参考基因的稳定性，筛选出各时期最稳定的复合参考基因。此外，使用产前地塞米松暴露（PDE）的病理模型来验证所选复合参考基因的稳定性和可靠性。结果表明，两个参考基因作为复合参考基因进行归一化是最优的。宫内期可选择SDHA和TBP作为复合参照基因，而在PW6和PW12可选择YWHAZ和GAPDH，参照基因的选择无明显性别差异。上述复合参考基因在PDE模型中保持稳定，可以使实验结果的统计显着性更加显着。综上所述，