Validation of reference genes for RT-qPCR analysis of CYP4T expression in crucian carp

Genetics and Molecular Biology

Fei Mo¹  Jie Zhao¹  Na Liu¹  Li-hua Cao¹  Shan-xiang Jiang¹ 

[1] ,Nanjing Agricultural University College of Veterinary Medicine Laboratory of Veterinary Pharmacology and ToxicologyNanjing Jiangsu Province ,China

关键词: gene expression;    quantitative RT-PCR;    housekeeping genes;    CYP4T;    crucian carp;   

DOI  :  10.1590/S1415-47572014000400005

来源: SciELO

【 摘 要 】

Reference genes are commonly used for normalization of target gene expression during RT-qPCR analysis. However, no housekeeping genes or reference genes have been identified to be stable across different tissue types or under different experimental conditions. To identify the most suitable reference genes for RT-qPCR analysis of target gene expression in the hepatopancreas of crucian carp (Carassius auratus) under various conditions (sex, age, water temperature, and drug treatments), seven reference genes, including beta actin (ACTB), beta-2 microglobulin (B2M), embryonic elongation factor-1 alpha (EEF1A), glyceraldehyde phosphate dehydrogenase (GAPDH), alpha tubulin (TUBA), ribosomal protein l8 (RPL8) and glucose-6-phosphate dehydrogenase (G6PDH), were evaluated in this study. The stability and ranking of gene expression were analyzed using three different statistical programs: GeNorm, Normfinder and Bestkeeper. The expression errors associated with selection of the genes were assessed by the relative quantity of CYP4T. The results indicated that all the seven genes exhibited variability under the experimental conditions of this research, and the combination of ACTB/TUBA/EEF1A or of ACTB/EEF1A was the best candidate that raised the accuracy of quantitative analysis of gene expression. The findings highlighted the importance of validation of housekeeping genes for research on gene expression under different conditions of experiment and species.

【 授权许可】

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