植物学报 ›› 2018, Vol. 53 ›› Issue (6): 829-839.DOI: 10.11983/CBB18003

• 技术方法 • 上一篇    下一篇

黄梁木实时荧光定量PCR分析中内参基因的选择

张登, 李景剑, 张梦洁, 包钰韬, 杨霄, 徐武云, 欧阳昆唏, 陈晓阳*()   

  1. 华南农业大学林学与风景园林学院, 广东省森林植物种质创新与利用重点实验室, 广州 510642
  • 收稿日期:2018-01-03 出版日期:2018-11-01 发布日期:2018-12-05
  • 通讯作者: 陈晓阳
  • 作者简介:作者简介:白克智, 1959年开始在中国科学院植物研究所工作, 先后任助理研究员、研究员, 长期从事植物生长发育及其调控的研究。1986年,其主持的“满江红生物学特性研究”荣获中国科学院科技进步二等奖。曾任《植物生理学报》编委、《植物学报》常务编委、中国植物生长调节剂协会主任等职。
  • 基金资助:
    国家自然科学基金(No.31600525)和广东省科技计划(No.2017B020201008)

Selection and Validation of Reference Genes for Quantitative RT-PCR Analysis in Neolamarckia cadamba

Zhang Deng, Li Jingjian, Zhang Mengjie, Bao Yutao, Yang Xiao, Xu Wuyun, Ouyang Kunxi, Chen Xiaoyang*()   

  1. Guangdong Key Laboratory for Innovative Development and Utilization of Forest Plant Germplasm, College of Forestry and Landscape Architecture, South China Agricultural University, Guangzhou 510642, China
  • Received:2018-01-03 Online:2018-11-01 Published:2018-12-05
  • Contact: Chen Xiaoyang

摘要:

为筛选黄梁木(Neolamarckia cadamba)实时定量PCR最佳内参基因, 该研究以黄梁木的根、芽、叶、花、果、皮及形成层为材料, 利用RT-qPCR技术对ACTCACCYPEF1α等21个管家基因家族43个候选内参基因进行表达量分析, 并利用geNorm、NormFinder和BestKeeper软件进行内参基因稳定性分析。geNorm的分析结果显示, UPL基因的稳定性最高(M=0.443), UBQ基因的稳定性最低(M=2.859); NormFinder的分析结果显示, UPL基因的稳定性最高(E=0.223), UBQ基因的稳定性最低(M=4.759); BestKeeper分析显示, UPL基因的标准偏差(SD=0.513)最低。研究结果表明, UPL基因作为内参基因稳定性最高, UBQ基因的稳定性最低。因此可以选择UPL基因作为黄梁木不同组织中RT-qPCR定量分析的内参基因。

关键词: 黄梁木, 实时荧光定量PCR, 内参基因

Abstract:

In this study, root, shoot, leaf, flower, fruit, peel and cambium tissue of Neolamarckia cadamba was sampled to analyze the expression of 43 candidate reference genes in 21 housekeeping gene families, such as ACT, CAC, CYP, and EF1α, by RT-qPCR. The software geNorm, NormFinder and BestKeeper were used to analyze expression stability of these candidate reference genes in the seven different tissues. geNorm analysis revealed that the stability of the UPL gene was the highest (M=0.443) and the stability of UBQ was the lowest (M=2.859). NormFinder analysis revealed that the stability of UPL was the highest (E=0.223), UBQ was the lowest (M=4.759). BestKeeper analysis revealed that the standard deviation of UPL (SD=0.513) was the lowest. These findings suggest that UPL has the highest stability, and UBQ has the lowest stability. UPL gene could be selected as an internal reference gene for analysis of gene expression among different tissues of N. cadamba.

Key words: Neolamarckia cadamba, real-time quantitative PCR, reference gene