芹菜肉桂酰辅酶A还原酶基因的克隆与表达分析

Cloning and the expression profiles analysis of an AgCCR gene encoding cinnamoyl-Co A reductase from different cultivars of Apium graveolens

[目的]肉桂酰辅酶A还原酶CCR(cinnamoyl-Co A reductase)在植物木质素代谢中起重要的作用,能够将羟基肉桂酸还原生成肉桂醛。本文目的是研究不同芹菜中肉桂酰辅酶A还原酶基因AgCCR在芹菜中组织表达和非生物胁迫过程中的响应情况。[方法]以2个芹菜品种‘六合黄心芹’和‘文图拉’为材料,分别克隆出编码肉桂酰辅酶A还原酶的基因AgCCR。利用荧光定量PCR分析不同芹菜组织和2种芹菜在高温、低温、干旱和盐胁迫下AgCCR的表达情况。[结果]序列分析表明,‘六合黄心芹’和‘文图拉’中AgCCR基因均含有1个长度为1 014 bp的开放阅读框,编码337个氨基酸,二者的核苷酸位点有3个位点的差异,编码的氨基酸有1个位点的差异。对2个芹菜中的AgCCR基因进行多重比对并绘制进化树,发现其编码的氨基酸序列与其他15种植物的CCR蛋白同源性较高,氨基酸高度保守。荧光定量PCR(qRT-PCR)分析表明,芹菜AgCCR基因在‘六合黄心芹’和‘文图拉’叶中的表达水平较高,AgCCR基因在高温和低温下表达上调,在干旱处理下表达下调。[结论]芹菜AgCCR基因对多种非生物胁迫均有响应。特别是低温下该基因响应明显,表明AgCCR基因或许与芹菜抗寒机制相关。

[ Objectives ] Cinnamoyl-CoA reductase can catalyze three kinds of hydroxy cinnamic acid CoA ester into cinnamic aldehyde, which plays an important role in the biosynthesis of lignin monomer. This study aims to explore the expression patterns of AgCCR gene in different tissues and various abiotic stresses in different celery cultivars. [ Methods]AgCCR gene was cloned from two celery cuhivars ＇ Liuhehuangxinqin＇ and＇ Ventura＇ ,respectively. Bioinformatic methods were used to analyze the sequence information,and qRT-PCR was performed to analyze the expression patterns of AgCCR gene in different tissues （root, petiole, leaf blade）and four abiotic stresses （cold,heat,drought and salt treatments） in celery. [ Results ] AgCCR gene of two cultivars contained an open reading frame with the length of 1 014 bp, which encoded 337 amino acids. The sequence analysis revealed that AgCCR gene from two cultivars had three nucleotide sites and one amino acid difference. Multiple alignment and phylogenetic tree showed that the amino acid sequences encoded by AgCCR gene from two cnltivars were highly homologous with CCR proteins from other 15 species, and these proteins were highly conserved, qRT-PCR analysis demonstrated that the AgCCR gene mainly expressed in leaf and responded to abiotic stresses. AgCCR gene was greatly upregulated after high and low temperature treatments and downregnlated after drought treatment in two celery cultivars. [ Conclusions ] AgCCR gene in celery was related to abiotic stress, and it responded obviously under low temperature treatment. The AgCCR gene may relate to mechanism of cold resistance in celery.