中国小麦花叶病毒(CWMV)侵染条件下小麦内参基因的选择

Selection of reference genes in wheat infected by Chinese wheat mosaic virus( CWMV)

实时定量PCR(qRT-PCR)是分析功能基因表达水平的常用方法之一,qRT-PCR数据统计分析离不开合适内参基因的选择。为了选择中国小麦花叶病毒(Chinese wheat mosaic virus,CWMV)侵染条件下的小麦内参基因,本实验通过PCR扩增效率和扩增特异性分析,从10个持家基因中选择8个候选基因,然后以接种CWMV的小麦样品为材料,利用qRT-PCR技术进一步检测上述8个基因在CWMV侵染条件下小麦样品中的时空表达特性。基于这些数据,通过ge Norm、Norm Finder程序分析,结果表明,2个小麦基因(即26S和CDC)在CWMV侵染前后以及不同组织中表达最为稳定,26S和CDC组合可选作CWMV与小麦互作过程中功能基因表达分析的内参基因。

Quantitative PCR （qRT-PCR） has been widely applied to the expression analysis of functional genes. The selection of appropriate reference genes is required for the statistical analysis of qRT-PCR data. To select the suitable reference genes in wheat infected by Chinese wheat mosaic virus （CWMV）, 10 housekeeping genes were screened by calculations of PCR efficiencies and melting curve analysis, suggesting that 8 out of 10 genes could be used as candidate reference genes. And then their temporal and spatial expression patterns were investigated by qRT-PCR in the different tissues and at the different stages of wheat inoculated with CWMV. These quantitative data were further evaluated by the computer programs geNorm and NormFinder in this study. The analysis results consistently revealed that two genes, 26 S and CDC , were stably expressed in all tested tissues and CWMV-infected wheat samples, indicating that the combination of 26 S and CDC could be selected as reference genes for expression analysis of functional gene during the interaction between CWMV and wheat.