鸭茅根组织实时定量PCR分析中候选内参基因的筛选  

Candidate Reference Gene Selection for Quantitative RT-PCR Normalization in Orchardgrass(Dactylis glomerata L.) Root Tissue

作　　者：蒋晓梅[1] 严海东[1] 张新全[1] 张瑜[1] 黄琳凯[1] 

机构地区：[1]四川农业大学草业科学系,四川雅安625014

出　　处：《草地学报》2014年第4期847-853,共7页Acta Agrestia Sinica

基　　金：国家973计划(2014CB138705);国家现代牧草产业技术体系(CARS-35-05);国家级大学生创新计划(201310626002)资助

摘　　要：实时荧光定量PCR(qRT-PCR)技术已成为研究基因表达的重要技术之一。以鸭茅(Dactylis glomerata)根组织为材料,利用qRT-PCR技术检测ACTIN(肌动蛋白酶)、TCTP(翻译控制肿瘤蛋白酶)、UBC(泛素结合酶)、ZTL(ZTL蛋白酶)、GAPDH(甘油醛-3-磷酸-脱氢酶)和SAMDC(腺苷甲硫氨酸脱羧酶)6个候选内参基因在不同非生物胁迫环境下的表达情况,并运用Genorm 3.5,Delta-Ct method,NormFinder 0.953,Bestkeeper 1.0和RefFinder软件分析各候选内参基因在鸭茅根组织中的表达稳定性。结果表明:6个候选内参基因综合表达稳定性从高到低排序为ACTIN,TCTP,GAPDH,UBC,ZTL,SAMDC;在干旱胁迫、盐胁迫、涝胁迫、热胁迫与ABA处理条件下,表达稳定性最高的基因是ACTIN;在涝胁迫条件下,表达稳定性最高的基因是TCTP;在不同的胁迫条件下,表达稳定性最低的基因是SAMDC。综合分析可知,基因ACTIN和TCTP最适合作为鸭茅非生物胁迫研究的候选内参基因,这可为鸭茅非生物胁迫下各种基因表达研究奠定基础。qRT-PCR has become an important technology to research gene expression.Orchardgrass root tissues were used as materials in this study.The mRNA expression stabilities of six candidate reference genes including actin （ACTIN）,translationally controlled tumor protein （TCTP）,ubiquitin conjugating enzyme （UBC）,zeitlupe （ZTL）,glyceraldehyde 3 phosphate dehydrogenase （GAPDH） and S adenosylmethionine decarboxylase （SAMD） were detected by qRT PCR technology under different abiotic treatment conditions.Delta-Ct method,Genorm （ver.3.5）,Bestkeeper （ver.1.0）,NormFinder （ver.0.953） and RefFinder were used to analyze the expressing stabilities of candidate reference genes in orchardgrass root tissues.The recommended comprehensive ranking of six candidate reference genes from better to average was as follows：ACTIN,TCTP,UBC,ZTL,GAPDH and SAMD.Under drought,salt,waterlogging,heat and ABA treatment conditions,ACTIN was the most suitable gene comparatively.Under waterlogging stress,TCTP was the best gene.SAMD was the least stable reference gene under different abiotic stresses.The results of comprehensive analysis indicated that ACTIN and TCTP were the most suitable candidate reference genes for orchardgrass under various abiotic stresses.This study laid the foundation for the expression analyses of other genes in orchardgrass under various abiotic stresses.

关 键 词：鸭茅 实时荧光定量PCR 非生物胁迫 候选内参基因 根组织 

分 类 号：Q943.2[生物学—植物学]