《植物生理学报》 2015, 51(11): 2031-2038

松叶猪毛菜干旱胁迫下实时定量PCR内参基因的筛选

闻志彬¹, 张明理^1,2,*

¹中国科学院新疆生态与地理研究所, 中国科学院干旱区生物地理与生物资源重点实验室, 乌鲁木齐830011; ²中国科学院植物研究所, 北京100093

通信作者：张明理；E-mail: zhangml@ibcas.ac.cn；Tel: 0991-7823151

摘　要：

实时荧光定量PCR (qRT-PCR)是研究植物基因定量表达的常用手段之一, 选择合适的内参基因是获得可信定量结果的前提。本文以不同干旱胁迫处理下松叶猪毛菜的叶片为材料, 应用qRT-PCR技术分析5个常用候选管家基因(18S rRNA、α-tubulin、β-actin、EF1-α和GAPDH)的表达变化, 利用geNorm和NormFinder软件筛选出在干旱胁迫下表达最稳定的内参基因。结果表明, 以不同干旱胁迫处理的所有样品作为总样品池进行内参基因筛选时, 候选内参基因表达稳定性顺序依次是β-actin>α-tubulin>GAPDH>EF1-α>18S rRNA。对同一干旱处理的样品进行内参基因筛选时, 5个候选内参基因表达稳定性存在明显差异, 其中β-actin表达稳定度最高。因此, β-actin是研究松叶猪毛菜干旱胁迫下基因表达最合适的内参基因。

关键词：松叶猪毛菜; 实时荧光定量PCR; 内参基因; 干旱胁迫; 藜科

收稿：2015-09-15   修定：2015-11-09

资助：国家自然科学基金(31300217)和中国科学院西部之光项目(XBBS201209)。
致谢 中国科学院新疆生态与地理研究所研究生徐喆、张杰以及新疆大学干旱生态环境研究所的李岩博士在样品采集中给予帮助。

Reference Gene Selection for Real-Time Quantitative PCR in Salsola laricifolia under Soil Drought Stress

WEN Zhi-Bin¹, ZHANG Ming-Li^1,2,*

¹Xinjiang Institute of Ecology and Geography, Chinese Academy of Sciences, Key Laboratory of Biogeography and Bioresource in Arid Land, Urumqi 830011, China; ²Institute of Botany, Chinese Academy of Sciences, Beijing 100093, China

Corresponding author: ZHANG Ming-Li; E-mail: zhangml@ibcas.ac.cn; Tel: 0991-7823151

Abstract:

Real-time quantitative PCR (qRT-PCR) is one of the common technologies used for gene expression analysis. Selection of a suitable reference gene is a prerequisite to obtain the reliable results. In this study, five housekeeping genes including ribosomal RNA 18S (18S rRNA), α-tubulin, β-actin, elongation factor 1 alpha (EF1-α) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH) of the desert plant, Salsola laricifolia, were selected as candidate reference genes in the qRT-PCR, and two softwares, geNorm and NormFinder, were used to evaluate their expression stabilities in the leaves of S. laricifolia under soil drought stress in order to obtain a reliable reference gene. The results showed that there were significant differences among the five candidate reference genes under different soil drought stress treatments. And the order of expression stability was β-actin>α-tubulin>GAPDH>EF1-α>18S rRNA. While this order of expression stability was different under the same soil drought stress treatment, but the expression stability of β-actin was highest. In a word, β-actin was the most stable gene for analysis gene expression in S. lariciF_olia under soil drought stress.

Key words: Salsola laricifolia; real-time quantitative PCR; reference gene; soil drought stress; Chenopodiaceae