CAJ | 학술논문

提高实时荧光定量PCR(Real-time quantitative PCR, qRT-PCR)准确性的先决条件是选择表达稳定性较高的内参基因。本研究以多年生黑麦草(Lolium perenne L.)为材料，利用qRT-PCR技术检测翻译起始因子4A(eukaryotic initiation factor 4 alpha, eIF4A)、TNF结合蛋白Ⅰ(Tat binding protein-1, TBP-1)、泛素连接酶(ubiquitin-conjugating enzyme, E2)、多聚泛素酶基因(ubiquitin, UBQ)、生物钟受体蛋白(zeitlupe, ZTL)和甘油醛三磷酸脱氢酶(glyceraldehyde-3-phosphate dehydrogenase, GAPDH)6个候选内参基因在不同非生物胁迫下的表达情况。运用geNorm(ver.3.5)、Normfinder(ver.0.953)、BestKeeper(ver.1.0)、Delta Ct和RefFinder软件综合分析表明，在干旱、盐、热胁迫及ABA处理下，eIF4A基因表达稳定性最高；在涝胁迫下，UBQ基因表达稳定性最高。因此，eIF4A和UBQ候选基因可作为不同胁迫下的内参基因，为进一步开展多年生黑麦草基因表达的定量研究了提供了一定的理论依据。
제고실시형광정량PCR(Real-time quantitative PCR, qRT-PCR)준학성적선결조건시선택표체은정성교고적내삼기인。본연구이다년생흑맥초(Lolium perenne L.)위재료，이용qRT-PCR기술검측번역기시인자4A(eukaryotic initiation factor 4 alpha, eIF4A)、TNF결합단백Ⅰ(Tat binding protein-1, TBP-1)、범소련접매(ubiquitin-conjugating enzyme, E2)、다취범소매기인(ubiquitin, UBQ)、생물종수체단백(zeitlupe, ZTL)화감유철삼린산탈경매(glyceraldehyde-3-phosphate dehydrogenase, GAPDH)6개후선내삼기인재불동비생물협박하적표체정황。운용geNorm(ver.3.5)、Normfinder(ver.0.953)、BestKeeper(ver.1.0)、Delta Ct화RefFinder연건종합분석표명，재간한、염、열협박급ABA처리하，eIF4A기인표체은정성최고；재로협박하，UBQ기인표체은정성최고。인차，eIF4A화UBQ후선기인가작위불동협박하적내삼기인，위진일보개전다년생흑맥초기인표체적정량연구료제공료일정적이론의거。
The selection of high stablely expressed reference genes is an important prerequisite for improving the accuracy of Real-time quantitative PCR (qRT-PCR). We investigated the expression of 6 candidate reference genes including eukaryotic initiation factor 4 alpha (eIF4A), Ti-binding peptide-1 (TBP-1), ubiquitin-conjugating enzyme (E2), ubiquitin (UBQ), zeitlupe (ZTL) and glyceraldehyde-3-phosphate dehydrogenase (GAPDH) under various abiotic stresses in perennial ryegrass (Lolium perenne L.). The analysis with geNorm (ver. 3.5), Normfinder (ver. 0.953), BestKeeper (ver. 1.0), Delta Ct and RefFinder revealed that under drought, salt, heat and ABA stresses, eIF4A gene was the most stable reference gene;under waterlogging stress, UBQ gene was the most stable reference gene. As a result, eIF4A and UBQ genes were selected as reference genes for perennial ryegrass under different abiotic stresses, which provides a foundation for gene expression investigation of perennial ryegrass.