| Summary: | <i>Kengyilia</i> is a newly established genus. Most species in this genus survive in hash environment, which might be an indicator of an acquirement of stress resistance genes and the potential for molecular breeding in <i>Triticeae</i> species. Quantitative real-time PCR (qRT-PCR) is a widely used technique with varied sensitivity heavily dependent on the optimal level of the reference genes. <i>K. melanthera</i> is a typical psammophyte species which has high drought resistance. The reference genes of <i>K. melanthera</i> are not yet reported. This study aims to evaluate the expression stability of 14 candidate reference genes (<i>EF1A, GAPDH, ACT1, UBI, TUBB3, TIPRL, CACS, PPP2R1B, TUBA1A, EIF4A1, CYPA3, TCTP, ABCG11L,</i> and <i>FBXO6L</i>) under five treatments (drought, heat, cold, salt, and ABA) and find the most stable and suitable one even upon stressed conditions. The software NormFinder, GeNorm, BestKeeper, and RefFinder were used for data analysis. In general, the genes <i>CACS</i> and <i>PPP2R1B</i> are concluded to have the best overall performance under the various treatments. With the ABA treatment, <i>TCTP</i> and <i>TIPRL</i> show the best stability. <i>CACS</i> and <i>TCTP</i>, as well as <i>TIPRL</i> and <i>CYPA3</i>, were most stable under the treatments of cold and salt, respectively. <i>CACS</i> and <i>FBXO6L</i> were ranked the highest with the heat treatment and drought treatment, respectively. Finally, the Catalase-1 (<i>CAT1</i>) gene was used to verify the reliability of the above reference genes. Accordingly, <i>CAT1’s</i> expression pattern remained unchanged after normalization with stable reference genes. This study provides beneficial information about the stability and reliability of potential reference genes for qRT-PCR in <i>K. melanthera</i>.
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