适用于大豆实时荧光定量PCR分析的内参基因的筛选和验证  

Selection and validation of reference genes for quantitative RT-PCR analysis in soybean

作　　者：曾文韬 柴春月[1,2] 窦道龙[1] 

机构地区：[1]南京农业大学植物保护学院,江苏南京210095 [2]南阳师范学院生命科学与技术学院,河南南阳473061

出　　处：《南京农业大学学报》2015年第5期787-795,共9页Journal of Nanjing Agricultural University

基　　金：国家转基因生物新品种培育重大专项(2014ZX0800910B)

摘　　要：[目的]选择合适的内参基因是实时荧光定量PCR(qRT-PCR)研究的关键。大豆由于基因组复杂等原因,内参基因的选择尤为重要。[方法]结合实验室先前试验结果及相关文献,选择了10个持家基因(Cons4、ACT11、TUA、Cons15、ACT20、Cons7、CYP2、Cons6、Tubulin and ELF1B),依次用3个统计学软件Ge Norm、Norm Finder和Best Keeper进行综合分析,研究其在大豆不同组织和大豆疫霉侵染不同时间点根中的表达稳定性。[结果]Cons4、ACT11和TUA在大豆疫霉侵染过程中的大豆根中表达稳定性最好,而Tubulin和ELF1B在大豆的所有样品中表达稳定性都较差。[结论]Cons4、ACT11、TUA可以推荐为大豆疫霉侵染病程中大豆基因表达qRT-PCR分析的首选内参基因;Cons15、CYP2、Cons4可以推荐为研究苗期大豆不同组织中基因表达qRT-PCR分析的内参基因。[Objectives]Selection of suitable reference genes is a critical step in real-time quantitative PCR（ qRT-PCR） analysis.[Methods]According to the reported,we selected ten housekeeping soybean genes,including Cons4,ACT11,TUA,Cons15,ACT20,Cons7,CYP2,Cons6,Tubulin and ELF1 B,and analyzed their expression patterns in various tissues of soybean seedings and at different time ponits of soybean roots treated by Phytophthora sojae. The stability of the ten genes expression was analyzed using three available software packages,Ge Norm,Norm Finder and Best Keeper. [Results]The genes Cons4,ACT11 and TUA expression are the most stable during P. sojae infection stages. Two genes,Tubulin and ELF1 B,are not suitable for reference genes. [Conclusions]The genes Cons4,ACT11 and TUA were suitable to determine gene expressional profiles upon P. sojae infection; and Cons15,CYP2 and Cons4 could be used for various tissues of soybean seedings.

关 键 词：大豆 实时荧光定量PCR 内参基因 大豆疫霉诱导表达 

分 类 号：S432.1[农业科学—植物病理学]