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2025年4月4日 星期五
  2017, Vol. 25 Issue (8): 1356-1365    
  研究资源与技术改进 本期目录 | 过刊浏览 | 高级检索 |
紫云英实时荧光定量PCR分析中内参基因的选择
张贤1,王建红2,李增强1,曹凯1,庄俐1,徐昌旭1,曹卫东2
1. 浙江省农业科学院
2.
Reference Gene Selection for Real-time Quantitative PCR Normalization in Astragalus sinicus
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摘要 摘 要 在实时荧光定量PCR(qRT-PCR)分析中,选择合适的内参基因是保证检测结果准确性的先决条件。紫云英(Astragalus sinicus)基因组信息匮乏,内参基因的选择尤为重要。本研究筛选了7个持家基因GAPDH、Cons7、ELF1B、Tubulin、Ubiquitin E2、Actin A以及Actin 20,利用qRT-PCR 技术研究其的表达情况,并采用GeNorm、NormFinder和Best Keeper软件,分析其在紫云英不同组织(根、茎、叶、花和荚果)和不同非生物胁迫(干旱胁迫、涝胁迫、盐胁迫、脱落酸(abscisic acid, ABA)胁迫)条件下叶片中的表达稳定性。结果表明,7种内参基因在紫云英不同的组织器官和不同的胁迫处理分析中,稳定性表现不尽相同。Cons7 在盐胁迫和涝胁迫处理下最稳定,Tubulin、Actin 20和GAPDH分别在不同组织、ABA处理和干旱胁迫下表达最稳定。因此,采用qRT-PCR分析紫云英不同器官组织中的基因表达差异时,可选择Tubulin作为校正内参基因;而比较脱落酸处理、盐胁迫和涝胁迫处理时,可选择Cons7作为校正内参基因;比较干旱胁迫处理时,选择GAPDH作为校正内参基因。本研究结果为研究紫云英不同生理过程中目的基因表达提供了稳定可靠的内参基因。
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张贤
王建红
李增强
曹凯
庄俐
徐昌旭
曹卫东
关键词 紫云英实时荧光定量PCR内参基因筛选    
Abstract:Abstract In the qRT-PCR analysis, selection of suitable reference genes is a prerequisite for the guarantee of accurate result. As the lack of genome information, the reference gene selection is particularly important for Chinese milk vetch (Astragalus sinicus). In this study, expression of seven commonly used housekeeping genes such as GAPDH, Cons7, ELF1B, Tubulin, Ubiquitin E2, Actin A and Actin 20 were assessed by qPCR in different tissues (roots,stems,leaves, flowers and pods) and different treatment (drought stress, waterlogging stress, salt stress, abscisic acid (ABA) stress) of Astragalus sinicus from NingBo. To determine the expression stability of these genes, qRT-PCR data were determined by three software tools (GeNorm, NormFinder and BestKeeper). The results showed that the highest expression stability candidate reference genes of Chinese milk vetch had many differences in the various experimental conditions. Cons7 could express stably under salt stress and waterlogging stress, and Tubulin, Actin 20, GAPDH were top ranked under different tissues, ABA treatment and drought stress respectively. In different tissues, Tubulin could be the best housekeeping gene. Cons7 was considered as the most appropriate housekeeping gene of Chinese milk vetch among abscisic acid treatment, salt stress and flooding stress, while GAPDH could be chosen as the best housekeeping gene under drought stress. The reference genes selected in this study will be helpful for improving the quality of gene expression studies under various stresses in Chinese milk vetch
Key wordsAstragalus sinicus    qRT-PCR    Reference gene    Selection
收稿日期: 2017-02-10      出版日期: 2017-07-20
基金资助:浙江省自然科学基金;公益性行业(农业)科研专项
通讯作者: 张贤     E-mail: zhangxian0399@126.com
引用本文:   
张贤 王建红 李增强 曹凯 庄俐 徐昌旭 曹卫东. 紫云英实时荧光定量PCR分析中内参基因的选择 [J]. , 2017, 25(8): 1356-1365.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/     或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2017/V25/I8/1356
 
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