油菜柠檬酸合酶基因的克隆及在逆境下的表达

doi:10.3724/SP.J.1006.2009.00033

摘要/Abstract

摘要：

柠檬酸合酶是植物多种代谢途径的限速酶, 及代谢变化的标志酶。为了解油菜柠檬酸合酶基因的生理功能, 以甘蓝型油菜叶片cDNA为模板, 设计特异性引物, 获得一编码柠檬酸合酶基因的cDNA序列, 全长1 659 bp, ORF区含476个氨基酸残基, 序列比对显示其蛋白序列与拟南芥有较高的同源性(96.0%), 氨基末端含一线粒体靶信号。聚类分析表明, 油菜柠檬酸合酶基因与其他植物内该基因高度同源。对油菜幼苗进行植物生长调节物质、高温和低温、强光照和弱光照、盐、菌核病、干旱和水渍等处理, 采用半定量PCR法对油菜叶片柠檬酸合酶基因的表达模式进行检测, 发现在盐胁迫、暗光和强光的处理下, 柠檬酸合酶基因的表达基本没有变化；在水渍、干旱、IAA和6-BA胁迫下, 其表达有所升高, 但出现峰值的时间不同, ABA对表达模式的影响与IAA相反；感染菌核病后其表达降低；对GA₃的应答呈鞍型。对部分处理采用荧光定量PCR验证, 其结果与半定量PCR结果基本一致。

关键词: 油菜, 柠檬酸合酶, 表达模式, 胁迫

Abstract:

As a key enzyme of some metabolisms, citrate synthase shows a sign for metabolism. In order to investigate the function of citrate synthase gene, the cDNA encoding citrate synthase was cloned from rapeseed leaf by RT-PCR. It was 1 659 bp long and encoded a protein with 476 amino acids. The deduced protein sequence had a mitochondrial targeting signal in N-terminal, which was very similar to the citrate synthase in Arabidopsis thaliana (96.0%). Alignment analysis showed that citrate synthase gene had high homology in plants. Under different stresses, we tested the expression of citrate synthase gene in rapeseed leaf by using semi-quantitative PCR. The expression of citrate synthase gene had no obvious change in stresses of salt, dark, high illumination, while was increased at different time in treatments of water logging, drought, IAA, and 6-BA. Interestingly the effect of ABA was contrary to that of IAA. In the treatment of sclerotium blight, the expression of citrate synthase gene was depressed. There was a saddle curve of citrate synthase gene expression in the treatment of gibberellin. The results from real-time PCR of several treatments were mainly in accord with that from semi-quantitative PCR.

Key words: Rapeseed(Brassica napus L.), Citrate synthase, Expression pattern, Stress

童晋;詹高淼;王新发;刘贵华;华玮;王汉中. 油菜柠檬酸合酶基因的克隆及在逆境下的表达[J]. 作物学报, 2009, 35(1): 33-40.

TONG Jin,ZHAN Gao-Miao, WANG Xin-Fa,LIU Gui-Hua,HUA Wei,WANG Han-Zhong. Cloning of Citrate Synthase Gene in Rapeseed(Brassica napus L.) and Its Expression under Stresses[J]. Acta Agron Sin, 2009, 35(1): 33-40.

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