植物学报 ›› 2013, Vol. 48 ›› Issue (2): 184-191.DOI: 10.3724/SP.J.1259.2013.00184

• 技术方法 • 上一篇    下一篇

水稻虫害诱导相关基因实时定量PCR中内参基因的选择

李冉, 李建彩, 周国鑫, 娄永根*   

  1. 浙江大学昆虫科学研究所, 杭州 310029
  • 收稿日期:2012-05-22 修回日期:2012-11-02 出版日期:2013-03-01 发布日期:2013-04-07
  • 通讯作者: 娄永根
  • 基金资助:

    国家重点基础研究发展计划;国家青年科学基金项目

Validation of Rice Candidate Reference Genes for Herbivore-induced Quantitative Real-time PCR Analysis

Ran Li, Jiancai Li, Guoxin Zhou, Yonggen Lou*   

  1. Institute of Insect Sciences, Zhejiang University, Hangzhou 310029, China
  • Received:2012-05-22 Revised:2012-11-02 Online:2013-03-01 Published:2013-04-07
  • Contact: Yonggen Lou
  • Supported by:

    973

摘要: 实时定量PCR技术广泛应用于植物功能基因转录水平变化的研究, 选择合适的内参基因进行相对定量分析是实验结果准确的关键因素。通过分析5个常用的内参基因(eEF-1α18S rRNA25S rRNAActinUBQ5)在水稻(Oryza sativa)经过各种处理后表达的稳定性, 结果表明, 水稻经过机械损伤处理后eEF-1α基因的表达最稳定; 二化螟处理后25S rRNA基因的表达最为稳定; 稻纵卷叶螟处理后Actin基因的表达最稳定; 两种刺吸式口器昆虫褐飞虱和白背飞虱危害后, UBQ5基因的表达最稳定。同时, 利用OsHI-LOX基因在不同处理后的表达来评价这些内参基因。研究结果为水稻虫害诱导实时定量PCR分析中内参基因的选择提供了理论依据。

Abstract: Quantitative real-time PCR is popularly used to study gene transcription levels in plants. Validation of candidate reference genes is the key step to ensure relative quantification. Here, we selected 5 reference genes (eEF-1α, 18S rRNA, 25S rRNA, Actin, and UBQ5) and tested their expression stability in rice under various treatments. eEF-1α showed the most stable expression with mechanical wounding, 25S rRNA with stripped stem borer treatment, and Actin with leaf folder treatment. UBQ5 showed the most stable expression on treatment with 2 phloem-feeding herbivores, brown plant hopper and white-backed plant hopper. We validated the expression profiles of these candidate reference genes by studying that of OsHI-LOX. We offer an available reference for studying gene transcription levels with herbivore infestation in rice.