摘要
抗坏血酸过氧化物酶(APX)是一种在逆境条件下清除细胞内氧自由基、增强植物抗逆性的关键酶。本研究根据已发表植物抗坏血酸过氧化物酶基因APX的同源保守序列设计引物,采用RT-PCR扩增超强抗寒白菜型冬油菜陇油7号的DNA,获得APX基因开放阅读框,长度为753 bp,编码250个氨基酸残基,推导的氨基酸序列具有APX蛋白的典型特征。生物信息学分析显示,与已报道的大白菜的APX氨基酸序列同源性达99%,该酶蛋白具有高度的进化保守性,其保守序列属于植物的POD超家族,APX相对分子质量和理论等电点依次为27.7 kD和5.58;APX基因无信号肽,是一个亲水性蛋白,可推测其定位于细胞质中;二级结构预测表明陇油7号的APX是由不规则卷曲和α-螺旋组成的稳定蛋白。实时荧光定量PCR和酶活性分析显示,该基因表达和酶活性受低温胁迫诱导,表明该基因在白菜型冬油菜陇油7号适应低温胁迫过程中发挥重要作用。
Ascorbate peroxidase（APX） is a key enzyme which eliminates oxygen free radicals and increases plant resistance in adverse circumstances.The complete open reading frame of APX gene was cloned from an extremely low temperature（–32℃） resistant winter turnip rape（Brassica campestris L.） cultivar Longyou 7 using the method of reverse transcription PCR（RT-PCR）.The primers were designed according to the published APX cDNA sequences.The sequence of APX from B.campestris L.was 753 bp,encoding a protein of 250 amino acid residues with a predicted molecular weight of 27.7 kD and a theoretical pI of 5.58.Bioinformatics analysis showed that the predicted APX protein contained a conserved amino acid sequence corresponding to the plant peroxidase（POD） superfamily.The APX had no signal peptide and was a hydrophilic protein locating in cytoplasm.The prediction of the second structures indicated the APX was a steady protein with more random coils and α-helices.The expression analysis of APX gene using real time RT-PCR and the activity of APX in response to lower temperature showed that the APX could be induced and the activity of APX could be changed by lower temperature.In conclusion,APX gene might play a role in cold tolerance of the B.campestris L.cultivar Longyou 7.
出处
《作物学报》
CAS
CSCD
北大核心
2013年第8期1400-1408,共9页
Acta Agronomica Sinica
基金
国家高技术研究发展计划(863计划)项目(2011AA10A104)
国家公益性行业(农业)科研专项经费项目(200903002-04)
国家现代农业产业技术体系建设专项(CARS-13)资助
关键词
白菜型油菜
低温
APX基因克隆
表达分析
APX活性
Winter rape（Brassica compestris L.）
Lower temperature
APX gene cloning
Expression analysis
Activity of APX
