桉属植物内参基因的筛选及评估  

Selection and evaluation of internal reference genes in Eucalyptus species

作　　者：黄真池[1] 欧阳乐军[1,2] 张龙[1,2] 沙月娥[1,2] 曾富华[1] 

机构地区：[1]湛江师范学院生命科学与技术学院,广东湛江524048 [2]湖南农业大学生物科学与技术学院,湖南长沙410128

出　　处：《西北农林科技大学学报（自然科学版）》2013年第10期67-72,共6页Journal of Northwest A&F University(Natural Science Edition)

基　　金：广东省自然科学基金项目(10452404801004328;8152404801000011;S2012010008737);国家星火计划项目(2012GA780019;2011GA780067)

摘　　要：【目的】分析桉属植物候选内参基因的表达稳定性,筛选合适的内参基因,为深入研究桉属植物愈伤组织芽分化及体细胞胚胎发生过程中基因的表达变化提供参考。【方法】以尾巨桉、粗皮桉、尾叶桉的叶片及尾叶桉的绿色、红色和白色3种愈伤组织为材料,提取其总RNA后逆转录得cDNA;分别将桉属植物actin基因和拟南芥actin基因的GenBank序列输入Primer3Plus软件,设计Real-time quantitative PCR(qPCR)引物,同时选择RTEF和RARS为候选内参基因;以cDNA为模板进行qPCR,得4个候选内参基因的Ct值;并用RefFinder软件分析候选内参基因的表达稳定性。【结果】RefFinder软件分析结果显示,4个候选内参基因RTEF、RARS、EACT、AACT的稳定系数分别为1.189,1.861,2.828和3.224,基因表达稳定性由高到低依次为RTEF、RARS、EACT、AACT。【结论】以RTEF为内参基因分析桉属植物的基因表达变化较为理想。【Objective】The suitable internal reference gene was selected from 4 candidate internal reference genes by evaluating their expression stabilities.【Method】The total RNA extracted from 6 Eucalyptus tissues,including the Eucalyptus urophylla leaf,the Eucalyptus urophylla×Eucalyptus grandis leaf,the Eucalipu pellida leaf,and the green callus,the red callus and the white callus of Eucalyptus urophylla,was used as template to get cDNA by reverse transcription PCR.4 candidate internal reference genes,including actin of Eucalyptus （EACT AB505624.1）,actin of Arabidopsis （AACT NM_114519.2）,RTEF（EQ974103.1） and RARS（EQ973784.1） were used to perform the real-time quantitative PCR（qPCR）.Then,the Ct values of qPCR were analyzed by RefFinder software.【Result】The stability values of 4 candidate reference genes,RTEF,RARS,EACT and AACT,were 1.189,1.861,2.828 and 3.224,respectively.RTEF was the most stable gene,and AACT transcript showed maximum variation.【Conclusion】RTEF gene was suitable for gene expression analysis as internal reference gene in Eucalyptus species.

关 键 词：内参基因 桉属 愈伤组织 RefFinder软件 

分 类 号：S718.43[农业科学—林学]