At present, transcription analysis of gene expression commonly uses housekeeping genes as control for normalization. In this study, the expression levels of three housekeeping genes including GAPDH, β-actin, and 18S ...At present, transcription analysis of gene expression commonly uses housekeeping genes as control for normalization. In this study, the expression levels of three housekeeping genes including GAPDH, β-actin, and 18S rRNA in six tissues and five developmental stages of the Mandarin fish Siniperca chuatsi were assayed with quantitative real-time PCR （qPCR）. Differences in expression levels were analyzed using geNorm program. The results demonstrate that β-actin is the most stable gene at developmental stages and GAPDH is the most stable in different tissues. While 18S rRNA expression during development is differentially regulated, which indicates it is suitable as an internal control for gene expression normalization at the developmental level. Overall, the data suggest that the two most stable housekeeping genes are enough to accurately calibrate gene expression in S. chuatsi. The significance of this study provided convincing references and methodology for housekeeping gene selection and normalization in gene expression analysis with regular PCR or qPCR.展开更多
本研究分别以β-actin、18SrRNA和GAPDH为内参基因,采用实时荧光定量PCR对草鱼早期发育时期肌球蛋白重链(myosin heavy light,MYH)基因的mRNA表达量进行分析,并比较不同内参基因对MYH基因mRNA表达水平检测结果的准确性。研究结果表明,以...本研究分别以β-actin、18SrRNA和GAPDH为内参基因,采用实时荧光定量PCR对草鱼早期发育时期肌球蛋白重链(myosin heavy light,MYH)基因的mRNA表达量进行分析,并比较不同内参基因对MYH基因mRNA表达水平检测结果的准确性。研究结果表明,以β-actin和GAPDH作为内参,MYH基因mRNA表达水平完全一致,其表达量从原肠到仔鱼阶段逐次递增,仔鱼与原肠期阶段相比表达量差异显著;当采用18S rRNA作为内参时,MYH基因mRNA在发育阶段的表达量呈不稳定状态。因此,β-actin和GAPDH均可作为内参基因,用于草鱼早期发育中MYH基因mRNA的相对定量研究;而18S rRNA作为内参时,可能会对检测结果造成偏差。本研究不仅准确的揭示了草鱼MYH基因mRNA的表达特征,并且为荧光定量PCR技术在鱼类基因表达研究方面提供了有价值的参考。展开更多
基金国家自然科学基金(3077164430972263)Aid Program for Science and Technology Innovative Research Team in Higher Educational Instituions of Hunan Province
文摘At present, transcription analysis of gene expression commonly uses housekeeping genes as control for normalization. In this study, the expression levels of three housekeeping genes including GAPDH, β-actin, and 18S rRNA in six tissues and five developmental stages of the Mandarin fish Siniperca chuatsi were assayed with quantitative real-time PCR （qPCR）. Differences in expression levels were analyzed using geNorm program. The results demonstrate that β-actin is the most stable gene at developmental stages and GAPDH is the most stable in different tissues. While 18S rRNA expression during development is differentially regulated, which indicates it is suitable as an internal control for gene expression normalization at the developmental level. Overall, the data suggest that the two most stable housekeeping genes are enough to accurately calibrate gene expression in S. chuatsi. The significance of this study provided convincing references and methodology for housekeeping gene selection and normalization in gene expression analysis with regular PCR or qPCR.
文摘本研究分别以β-actin、18SrRNA和GAPDH为内参基因,采用实时荧光定量PCR对草鱼早期发育时期肌球蛋白重链(myosin heavy light,MYH)基因的mRNA表达量进行分析,并比较不同内参基因对MYH基因mRNA表达水平检测结果的准确性。研究结果表明,以β-actin和GAPDH作为内参,MYH基因mRNA表达水平完全一致,其表达量从原肠到仔鱼阶段逐次递增,仔鱼与原肠期阶段相比表达量差异显著;当采用18S rRNA作为内参时,MYH基因mRNA在发育阶段的表达量呈不稳定状态。因此,β-actin和GAPDH均可作为内参基因,用于草鱼早期发育中MYH基因mRNA的相对定量研究;而18S rRNA作为内参时,可能会对检测结果造成偏差。本研究不仅准确的揭示了草鱼MYH基因mRNA的表达特征,并且为荧光定量PCR技术在鱼类基因表达研究方面提供了有价值的参考。
基金This work was supported by the National 973 Program (No.2001CB109006), the Scientific Funds of the Educational Bureau of Hunan Province (No.04C378), and the Ph.D Programs Foundation in Hunan Normal University (No. 24030610)
