小拟南芥实时荧光定量PCR(qRT-PCR)内参基因的筛选  

Screening of Reference Genes of Quantitative Real-Time PCR(qRT-PCR) in Arabidopsis pumila

作　　者：郑丽洁[1] 林军[1] 黄先忠[1] 

机构地区：[1]石河子大学生命科学学院,植物基因组学实验室,石河子832003

出　　处：《基因组学与应用生物学》2017年第2期774-783,共10页Genomics and Applied Biology

基　　金：国家自然科学基金项目(U1303302,31060149)资助

摘　　要：小拟南芥(Arabidopsis pumila),又名无苞芥(Olimarabidopsis pumila),为拟南芥的近缘物种。选择合适的内参基因对于提高实时荧光定量PCR(quantitative real-time PCR,qRT-PCR)分析基因表达的准确性至关重要。本研究在分析小拟南芥叶片转录组数据的基础上,筛选了12个表达稳定的候选内参基因,包括肌动蛋白1(actin-1,ACT-1)、α-微管蛋白1(α-tubulin,TUA1)、β-微管蛋白6(β-tubulin,TUB6)、翻译起始因子(translation initiation factor 2,IF2)、延伸因子1-α(elongation factor 1-alpha,EF1-α)、聚泛素14(polyubiquitin 14,UBQ14)、UBQ9、泛素连接酶35(ubiquitin-conjugating enzyme 35,UBC35)、转录起始因子1(transcription initiation factor,HAF1)、FK506结合蛋白62(FK506 binding protein 62,FKBP62)、miR319a和醛脱氢酶(aldehyde dehydrogenase,ALDH)。我们分别利用Norm Finder、Genorm和Bestkeeper软件分析了这12个基因在dH_2O、NaCl处理下及小拟南芥不同组织中的表达稳定性。研究表明,在dH_2O处理条件下表达稳定的基因为TUB6和FKBP62,在Na Cl处理条件下为EF1-α和IF2,在组织器官中为HAF1,这些基因适合作为相应试验条件下小拟南芥的内参基因。本研究为小拟南芥基因表达分析提供了可靠的内参基因,具有重要的参考价值。Arabidopsis pumila （synonym： Olimarabidopsis pumila）, is a close relative ofArabidopsis thaliana. In order to improve the accuracy of the analysis ofgene expression using quantitative Real-time PCR （qRT-PCR）, the selection of appropriate reference genes is very important. In the present study, 12 candidate genes were selected basing on the transcriptome data, which including actin I （A CT-I）, α-tubulin 1 （TUA1）, β-tubulin 6 （TUB6）, translation initiation factor 2 （IF2）, elongation factor 1-α （EFI-α）, polyubiquitin 14 （UBQ14）, UBQ9, ubiquitin conjugating enzyme 35 （UBC35）, transcription initiation factor 1 （HAF1）, FK506 binding protein 62 （FKBP62）, microRNA （miR319a） and aldehyde dehydrogenase （A LDH）. Their stability of expression under dH20 treatment, NaCl treatment and in different tissues ofA rabidopsis pumila were analyzed by using NormFinder, Genorm and Bestkeeper soft-ware respectively. The research showed that the expression of TUB6 and FKBP62 were the most stable in dH2O-treated samples, while in NaCl-treated samples were EFI-α and IF2 and in different tissues was HA F1, they were suitable for expression analysis of genes in A. pumila under dH2O, NaCl and in different tissues. The study provides reliable reference genes in expression analysis ofA. pumila genes, which has important reference value.

关 键 词：小拟南芥 无苞芥 盐胁迫 实时荧光定量PCR 内参基因 

分 类 号：Q943.2[生物学—植物学]