《植物生理学报》 2016, 52(7): 1059-1065

橡胶草实时荧光定量PCR内参基因评价

覃碧¹, 潘敏², 余海洋², 杨玉双¹, 甘霖¹, 刘实忠^1,*

¹中国热带农业科学院橡胶研究所, 海南儋州571737; ²海南大学环境与植物保护学院, 海口570228

通信作者：刘实忠；E-mail: sz_liu@126.com

摘　要：

橡胶草(Taraxacum kok-saghyz)是一种重要的产胶作物。为了筛选出橡胶草稳定可靠的内参基因用于实时荧光定量PCR (qRT-PCR)分析, 本研究从已公布的橡胶草转录组数据中选取18S rRNA、EIF4α、tubulin、β-actin、GAPDH和40S ribosomal protein (40SRP)作为候选基因, 通过qRT-PCR检测, 并利用geNorm、NormFinder、BestKeeper等软件对这6个候选内参基因在不同橡胶草种质、不同组织以及机械伤害和乙烯利处理后不同时间下的表达稳定性进行分析, 结果发现6个候选内参基因的表达稳定性各异, GAPDH和β-actin基因是不同种质及其不同组织以及不同处理中表达最稳定的内参基因。本研究结果为橡胶草基因表达分析提供了稳定可靠的内参基因。

关键词：橡胶草; 内参基因; 实时荧光定量PCR

收稿：2016-04-22   修定：2016-07-04

资助：中国热带农业科学院橡胶研究所基本科研业务费专项(1630022015005)、山东玲珑轮胎有限责任公司横向课题(H2012177)和国家自然科学基金(31300503)。

Evaluation of reference genes for quantitative real-time PCR expression studies in Taraxacum kok-saghyz Rodin

QIN Bi¹, PAN Min², YU Hai-Yang², YANG Yu-Shuang¹, GAN Lin¹, LIU Shi-Zhong^1,*

¹Rubber Research Institute, Chinese Academy of Tropical Agricultural Sciences, Danzhou, Hainan 571737, China; ²College of Environment and Plant Protection, Hainan University, Haikou 570228, China

Corresponding author: LIU Shi-Zhong; E-mail: sz_liu@126.com

Abstract:

Taraxacum kok-saghyz (TKS) is one of the most important rubber-producing crops. To screen stable and reliable reference genes for quantitative real-time PCR (qRT-PCR) analysis in TKS, 6 housekeeping genes from the public transcriptome data of TKS, including 18S rRNA, EIF4α, tubulin, β-actin, GAPDH, and 40S ribosomal protein (40SRP), were selected as candidate reference genes for analysis in this study. The expression stabilities of the 6 genes were analyzed by geNorm, NormFinder, and BestKeeper, respectively, in different TKS accessions and tissues, and under mechanical wounding and ethephon treamtents. This result indicates that the expression stability varied greatly between these genes. GAPDH and β-actin had the highest stabilities across different accessions and tissues, and under different treatments. The results in this study provide stable and reliable reference genes for gene expression analysis in TKS.

Key words: Taraxacum kok-saghyz; reference gene; quantitative real-time PCR