棉花纤维发育和体细胞胚发生过程中实时定量PCR内对照基因的筛选

被引：31

作者：

涂礼莉

张献龙

刘迪秋

金双侠

曹景林

朱龙付

邓锋林

谭家福

张存斌

机构：

[1] 华中农业大学作物遗传改良国家重点实验室

来源：

科学通报 | 2007年 / 20期

关键词：

棉花; 纤维发育; 持家基因; 内对照; 实时定量PCR(qRT-RCR); 体细胞胚胎发生;

D O I：

暂无

中图分类号：

S562 [棉];

学科分类号：

摘要：

实时定量PCR技术(qRT-PCR)能够快速地对多个样品进行基因表达分析,因此已成为芯片和微点阵数据验证的常用手段.为了得到更精确的数据,通常需要一个和多个内对照基因来平衡化qRT-PCR数据.目前一般选择持家基因(housekeeping gene)作为内对照,但很多持家基因的表达水平随着环境的改变而改变.在棉花纤维发育和体细胞胚发生机制研究过程中已经产生大量芯片和微点阵数据,但目前在采用实时定量PCR验证这些数据时都只用了一个持家基因来平衡化数据.为了验证其可靠性,本研究根据常用的持家基因(18S rRNA,Histone3,UBQ7,Actin,Cyclophilin,Gbpolyubiquitin-1和Gbpolyubi-quitin-2)设计了7对引物,用相对的绝对定量法验证了在棉花不同组织和不同发育阶段(21个样品)表达水平的稳定性,发现在纤维发育的后期(17 DPA(day post anthesis)以后),这些基因的表达都下调,而纤维发育后期特异表达基因AGP的表达从15DPA开始到27DPA一直都呈上升趋势.因此对于纤维系列特别是纤维发育后期基因的qRT-PCR更好的选择是相对的绝对定量.而对于非纤维系列,这些持家基因的表达水平相对纤维系列变化幅度较小,但为了得到更精确的表达数据,应该同时用Histone3,UBQ7和Gbpolyubiquitin-1一起来平衡化qRT-PCR数据.

引用

页码：2379 / 2385

页数：7

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