珙桐实时定量PCR内参基因的筛选及稳定性评价

任锐, 戴鹏辉, 李萌, 刘志明, 曹福祥*
中南林业科技大学生命科学与技术学院, 长沙 410004

通信作者:曹福祥;E-mail: csfucao@163.com

摘 要:

实时荧光定量PCR (qPCR)技术被广泛应用于基因表达分析, 选用合适的内参基因是运用该技术准确分析目标基因表达变化的前提。本研究以珙桐不同器官为材料, 采用qPCR技术分析了珙桐中6个管家基因(ACT7eIFEF1aGAPDHβ-TUB18S rRNA)及2个新内参基因(CACTIP41), 共8个候选内参基因的表达情况, 并利用geNorm、NormFinder和Best-Keeper3种软件对候选内参基因的表达稳定性进行了评价。结果表明, CACACT7在珙桐营养器官与生殖器官中均表达稳定, 且新内参基因CAC的表达稳定性优于ACT7; GAPDHEF1a18S rRNATIP41在珙桐营养器官与生殖器官中均表达不稳定, 尤其是GAPDH18S rRNA的表达稳定性更差; β-TUB在珙桐营养器官中表达更稳定, 而eIF在珙桐生殖器官中表达更稳定。

关键词:实时定量PCR; 内参基因; CAC基因; 珙桐

收稿:2016-08-03   修定:2016-09-18

资助:湖南省百人计划(112-0991)与中南林业科技大学博士后科研基金(0490016)。

Selection and stability evaluation of reference genes for real-time quantitative PCR in dove tree (Davidia involucrata)

REN Rui, DAI Peng-Hui, LI Meng, LIU Zhi-Ming, CAO Fu-Xiang*
College of Life Science and Technology, Central South University of Forestry and Technology, Changsha 410004, China

Corresponding author: CAO Fu-Xiang; E-mail: csfucao@163.com

Abstract:

Real-time quantitative PCR (qPCR) has been widely used in gene expression analysis. Selection of suitable reference genes is the first step in analyzing the expression variation of target genes accurately by qPCR. In the present study, we analyzed the expression patterns of eight candidate reference genes including six housekeeping genes (ACT7, eIF, EF1a, GAPDH, β-TUB and 18S rRNA) and two novel reference genes (BADH and TIP41) in different organs of dove tree (Davidia involucrata) by qPCR. Three softwares, geNorm, NormFinder and BestKeeper, were used to evaluate the expression stability of candidate reference genes. Gene CAC and ACT7 were identified as stable genes in both vegetative organs and reproductive organs. Moreover, the expression stability of novel reference gene CAC is better than that of ACT7. On the other hand, the gene expression quantity of GAPDH, EF1a, 18S rRNA and TIP41 showed unstable, especially the expression stability of GAPDH and 18S rRNA. β-TUB was proved to be stable reference gene for dove tree vegetative organs, while in reproductive organs eIF was recommended.

Key words: real-time quantitative PCR; reference gene; CAC gene; dove tree (Davidia involucrata)

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