Quantitative evaluation and selection of reference genes in a rat model of extended liver resection.
Quantitative evaluation and selection of reference genes in a rat model of extended liver resection.
复制标题
扩大肝切除大鼠模型中内参基因的定量评估和选择。
DOI：
10.1016/s1726-4901(08)70161-8
复制
发表时间：
2009
期刊：
影响因子：
--
通讯作者：
U. Dahmen
中科院分区：
文献类型：
--
作者：
W. Xing;M. Deng;Jinyan Zhang;Hai Huang;O. Dirsch;U. Dahmen
部分肝切除术（PHx）是研究肝再生的常用实验模型。实时定量PCR（qPCR）已成为选择基因表达谱的首选方法之一，以阐明肝脏功能和再生的调节。在PHx后1、2和7天，通过qPCR在70%和90%大鼠PHx模型中评价5种常用管家基因（HKG; Alb、UBC、Hprt、Ywhaz和GAPDH）的表达。我们建立了一个严格控制的qPCR程序，验证每个关键步骤，并通过线性回归和方差分析对基因表达稳定性进行统计学评价。我们的结果表明，HKG最适合于在扩展的90%PHx模型中评估基因表达是Hprt。当将相同量的来自所有样品的cDNA引入扩增反应中时，可以省略HKG的扩增。使用生物分析仪测定cDNA浓度被证明是一种简单且重复性好的方法。使用这种技术，响应于测试的实验条件的HKG的转录水平的潜在调节或管家基因的稳定性变得无关紧要。
Partial hepatectomy (PHx) is a frequently used experimental model for the study of liver regeneration. Real-time quantitative PCR (qPCR) has become the one of the methods of choice for expression profiling of selected genes in order to elucidate the regulation of liver function and regeneration. The expression of five commonly used housekeeping genes (HKGs; Alb, UBC, Hprt, Ywhaz, and GAPDH) were evaluated by qPCR in 70% and 90% rat PHx model at 1, 2, and 7 d after PHx. We set up a closely controlled qPCR procedure validating each critical step and the gene expression stability was statistically evaluated by linear regression and analysis of variance. Our results showed the HKG best suited for the evaluation of gene expression in the extended 90% PHx model is Hprt. The amplification of an HKG can be omitted when the same amount of cDNA from all samples is introduced into the amplification reaction. Determination of cDNA concentration employing the bioanalyzer proved to be an easy and reproducible approach. Using this technique the potential regulation of the transcription level of the HKG in response to the experimental condition tested or the stability of a housekeeping gene becomes irrelevant.