茭白荧光定量RT-PCR内参基因表达稳定性评价
Stability Evaluation of Reference Genes for Fluorescence Quantitative RT-PCR in Zizania latifolia
马佳琪 1刘彦承 1宋思晓 1缪旻珉 2张治平1
作者信息
- 1. 扬州大学园艺园林学院,扬州,225009
- 2. 扬州大学园艺园林学院,扬州,225009;扬州大学农学院,江苏省作物基因组学和分子育种重点实验室,扬州,225009
- 折叠
摘要
茭白(Zizanialatifolia)是禾本科多年生水生蔬菜,其产品器官是由菰(Zizania latifolia)与菰黑粉菌(Ustilago esculenta)共生膨大形成的可食用肉质茎.选取合适的茭白荧光定量RT-PCR内参基因,是研究菰和菰黑粉菌互作中茭白基因表达模式的基础.本研究以茭白不同时期的根、叶片、叶鞘、茎尖和肉质茎为研究材料,通过 RT-qPCR 技术检测 β-Actin、Actin3、β-ubulin、GAPDH、UBQ5、UBC1、PK、elf1α、eIF4a、SFua2a、18S rRNA等11个内参基因的表达情况,借助内参基因评价软件GeNorm、Normfinder和BestKeeper以及综合评价软件RefFinder对候选内参基因的表达稳定性进行分析,筛选出茭白适宜的内参基因,并使用茭白ZlADR1基因进行验证分析.结果表明,SFua2a和eIF4a,其次Actin3在所有样品中表达稳定性较好,适合作为茭白基因表达分析的内参基因.
Abstract
Zizania latifolia is a kind of perennial aquatic vegetable of Gramineae,and the edible organ was the swollen culm formed by the symbiotic of Zizania latifolia and Ustilago esculenta.The selection of suitable internal reference genes of fluorescence quantitative RT-PCR(RT-qPCR)is the basis of studying the gene expression pattern of the Z.latifolia in the interaction between host plants and fungus.In this study,roots,leaves,sheaths,stem apex and culm gall of Zizania latifolia at different stages were used as research materials.The expression levels of 11 reference genes such as β-Actin,Actin3,β-tubulin,GAPDH,UBQ5,UBC1,PK,elf1α,eIF4a,SFua2a and 18S rRNA were analyzed by RT-qPCR,and the expression stability of these reference genes were identified using the GeNorm,NormFinder,BestKeeper and RefFinder software,then the ZlADR1 gene was used for verification analysis to test the reliability of reference genes.The results showed that SFua2a and eIF4a followed by Actin3 were stably expressed in all samples and were the appropriate reference genes of Z.latifolia.
关键词
茭白(Zizania/latifolia)/内参基因/实时荧光定量PCRKey words
Zizania latifolia/Reference gene/Real-time quantitative PCR引用本文复制引用
基金项目
江苏省农业科技自主创新资金项目(CX203104)
江苏省作物基因组学和分子育种重点实验室开放课题项目(ML202006)
中国博士后科学基金项目(2015M580478)
出版年
2024
NETL
NSTL
万方数据