芍药油体钙蛋白基因PlClo克隆及表达分析

Cloning and expression analysis of herbaceous peony caleosin gene PlClo

以芍药(Paeonia lactiflora)种子RNA为模板,采用RACE (rapid amplification of cDNA ends)技术对芍药油体钙蛋白基因(PlClo)进行克隆及序列的生物信息学分析,并采用qRT-PCR (quantitative real-time PCR)技术对PlClo在芍药发育种子中的表达模式进行检测,结果表明:PlClo基因全长为908 bp,具有完整的开放阅读框720bp,共编码239个氨基酸,等电点为5.97,不具有信号肽,存在1个明显的跨膜结构域,属于稳定蛋白和亲水性蛋白;α-螺旋为该蛋白二级结构的主要构成部分,存在1个糖基化位点和39个潜在的磷酸化位点。芍药发育种子中PlClo基因的表达水平呈先上升后下降趋势,在花后60 d表达水平最高。这一结果为后续深入研究PlClo基因在芍药油脂稳定储存中的作用提供参考。

Taking RNA extracted from herbaceous peony(Paeonia lactiflora)seed as template,rapid amplification of cDNA ends(RACE)technique was used to clone caleosin gene(Clo),and its sequence was analyzed.Subsequently,quantitative real-time PCR(qRT-PCR)technique was used to detect its expression characterization in herbaceous peony developed seed.The results show that herbaceous peony Clo(PlClo)was 908 bp in full length,has a 720 bp open reading frame,and encodes 239 amino acids,with an isoelectric point of 5.97.PlClo has only one transmembrane domain,and does not have a signal peptide,belonging to the stable and hydrophilic protein.A-helixes were the main parts of the secondary structure,and one glycosylation site and 39 potential phosphorylation sites were found in PlClo.With the development of herbaceous peony seeds,the expression level of PlClo presented an initially increased trend and then downward trend,and had the highest expression level on the 60 th day after flowering.These results would lay the foundation for the further study of PlClo gene in grease-stable storage of herbaceous peony.