莲雾果实C4H基因的克隆及在NO处理下的表达分析

Cloning and Expression Analysis of Cinnamic Acid-4-hydroxylase from Wax Apple Fruit Under NO Treatment

以莲雾(Syzygium samarangense)果实RNA为模板,采用聚合酶链反应(polymerase chain reaction,PCR)技术对莲雾果实肉桂酸-4-羟基化酶(cinnamic acid-4-hydroxylase,C4H)基因进行克隆及生物信息学分析,并采用实时荧光定量PCR技术对采后莲雾果实C4H基因在一氧化氮(nitric oxide,NO)处理下的表达水平进行分析。结果表明:莲雾果实C4H基因长1849 bp,包含长为1518 bp的开放阅读框,编码505个氨基酸;预测的C4H蛋白质等电点为9.34,包括一个细胞色素P450和一个反式肉桂酸4-单加氧酶保守结构域,没有信号肽和跨膜结构域。莲雾果实贮藏期间,C4H基因的表达水平和木质素含量逐渐上升,且两者的相关系数达0.972,呈极显著的正相关。外源NO处理抑制了莲雾果实C4H表达水平和木质素的积累。

In the present study,cinnamic acid-4-hydroxylase gene(C4H)was cloned by polymerase chain reaction(PCR)technology from wax apple fruit(Syzygium samarangense),and bioinformatics analysis were performed.In addition,the C4H expression level under NO treatment was analyzed by real-time fluorescent quantitative PCR technology.The results showed that the cloned C4H gene(1849 bp)contained a 1518-bp open reading frame encoding a protein of 505-amino acid with a calculated isoelectric point of 9.34.There was no signal peptide and transmembrane domain,and shared conserved domains with other C4H family members,including cytochrome P450 and trans-cinnamate 4-monooxygenase domains.The expression level of C4H and lignin content increased gradually during storage and their correlation coefficient was 0.972.Besides,C4H expression level and lignin content of harvested wax apple fruit was inhibited significantly by NO.