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大豆不同发育时期及非生物胁迫下实时荧光定量PCR内参基因筛选

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为研究大豆在不同发育时期的不同组织、不同非生物胁迫下稳定表达的最适内参基因，以大豆"JN28"V1时期的根、茎、叶，R3、R4时期的豆荚，R5、R8时期的子粒，干旱、低温、盐、脱落酸(ABA)胁迫下的根和叶共15个样本为试验材料。选择8个候选内参基因(Actin，β-actin，CYP2。EF1-α，Fbox，GAPDH，TUB4，1 8SrRNA)进行实时荧光定量PCR检测，并分析8个候选内参基因表达的稳定性。利用geNorm、NormFinder、BestKeeper软件分析后，再经过RefFinder计算筛选出合适的内参基因。结果表明:4个软件的分析结果不同，以RefFinder综合分析结果显示，V1期的根和叶、R3期的豆荚、ABA胁迫下的根，最适的内参基因为Actin;V1期的茎、R4期的豆荚、R8期的子粒，最适内参基因为EF1-α;R5期的子粒、干旱胁迫下的叶，最适内参基因为Fbox;干旱胁迫下的根，最适的内参基因为CYP2;盐胁迫的根、ABA胁迫下的叶，最适内参基因为18SrRNA;低温胁迫下的叶，最适内参基因为β-actin;低温胁迫下的根，最适内参基因为EF1-α和18SrRNA;盐胁迫下的叶，最适内参基因为β-actin和18SrRNA。4个软件在全部组织及全部胁迫中综合分析结果均一致，在全部组织中最适内参基因为Actin，全部胁迫中最适内参基因为EF1-α。
Screening of Reference Genes Under Abiotic Stress and Different Development Stages of Soybean by Real-Time Fluorescence Quanti-tative PCR
In order to study the optimal reference genes stablely expressed in different tissues and under different abiotic stresses in different development stages of soybean,15 soybean samples were used as experimental materials,including roots and leaves in JN28 V1 stage,pods in R3 and R4 stages,grains in R5 and R8 stages,and roots and leaves under drought and low temperature salt ABA stress.Eight candidate reference genes(Actin,β-actin,CYP2,EF1-α,Fbox,GAPDH,TUB4,18SrRNA)were selected for real-time quantitative PCR detection,and the expression stability of the eight candidate reference genes was analyzed.After analysis with the geNorm NormFinder Best-Keeper software,the appropriate internal reference genes were selected by RefFinder calculation.The results showed that the analysis results of the 4 software were not the same.The comprehensive analy-sis results of RefFinder showed that the root of stage V1,leaf of stage V1 and root of stage R3 under ABA stress,the most suitable internal reference gene was Actin.EF1-α was the most suitable refer-ence gene for stem of stage V1 and pod of stage R4 and grain of stage R8.In R5 stage,Fbox was the most suitable reference gene for leaves under drought stress.The optimal reference gene for roots un-der drought stress was CYP2.The optimal reference gene for leaves under ABA stress was 18SrRNA.β-actin was the most suitable reference gene for leaves under low temperature stress.For roots under low temperature stress,the optimum reference gene were EF1-α and 18SrRNA.In leaves under salt stress,the most suitable reference genes were β-actin and 18SrRNA.The comprehensive analysis re-sults of the 4 software were consistent in all tissues and stresses.The most suitable reference gene was Actin in all tissues,and the most suitable reference gene was EF1-α in all stresses.

soybeanreal-time quantitative PCR(qRT-PCR)internal reference geneabiotic stressdevelopmental stage

王蕊、胡绍旺、刘金凤、张毓哲、姜玉石、刘思言、史安迪

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吉林农业大学生命科学学院,长春 130118

大豆 实时荧光定量PCR 内参基因 非生物胁迫 发育时期

国家重点研发计划子课题吉林省自然科学基金项目吉林省教育厅科学技术研究规划项目(2021)吉林农业大学本科生科技创新基金项目吉林农业大学本科生科技创新基金项目

2019YFD1002603-120190201168JC20192021

2024

10.13327/j.jjlau.2021.1219

吉林农业大学学报
吉林农业大学

吉林农业大学学报

CSTPCD北大核心
影响因子：1.014
ISSN：1000-5684
年,卷(期)：2024.46(1)
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