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| Selection and Evaluation of Reference Genes for qRT-PCR in Chinese Flowering Cabbage (Brassica campestris ssp. chinensis var. utilis) Under Heat Stress |
| PANG Qiang-Qiang, ZHU Bai-Bi, ZHOU Man, CHEN Yi-Song, WANG Ya-Qiang, SHI Guo-Bin, SUN Xiao-Dong* |
| Vegetable Research Institute, Hainan Academy of Agricultural Sciences/Key Laboratory of Genetic Resources Evaluation and Utilization of Tropical Fruits and Vegetables (Co-construction by Ministry and Province), Ministry of Agriculture and Rural Affairs/Hainan Provincial Engineering Research Center for Melon and Vegetable Breeding/Hainan Key Laboratory of Vegetable Biology, Haikou 571100, China |
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Abstract Choosing appropriate reference genes is crucial for enhancing the accuracy of real-time fluorescence quantitative PCR (qRT-PCR) analysis results. To screen for stable expression of internal reference genes suitable for heat stress in Chinese flowering cabbage (Brassica campestris ssp. chinensis var. utilis), this study selected 16 candidate reference genes based on RNA-seq data. The expression stability of these genes was evaluated using GeNorm, NormFinder, BestKeeper, and RefFinder at different times, tissues, and cultivars under heat stress.The results showed that casein kinaseⅠ (CKⅠ) exhibited optimal expression stability under different time and tissue treatments of heat stress; The gene with the most stable expression among different cultivars under heat stress was 28S ribosomal protein S27 (28SRPS27). The comprehensive analysis of all heat stress samples shows that CKⅠ has the best overall stability. The validation experiment confirmed that using CKⅠ as an internal reference gene effectively and accurately quantifies the expression changes of heat-responsive genes, indicating its suitability for standardizing qRT-PCR data related to heat stress in Chinese flowering cabbage. This study identified stable reference genes suitable for heat stress in Chinese flowering cabbage, and providing a basis for functional analysis of heat-resistant genes and molecular breeding in the future.
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Received: 21 November 2025
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Corresponding Authors:
* sxd1949@163.com
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