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2025年8月15日 星期五
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西伯利亚蓼Tau类谷胱甘肽转移酶基因的克隆与表达分析
刘明坤;刘关君;阎秀峰;刘昌财;魏志刚;刘桂丰;杨传平
东北林业大学林木遗传育种与生物技术教育部重点实验室,哈尔滨150040
Cloning and Expression Analysis of Tau Class Glutathione Transferase Gene from Polygonum sibiricum Laxm
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摘要 用RACE技术从西伯利亚蓼(Polygonum sibiricum Laxm.)中克隆了谷胱甘肽转移酶(PsGSTU1)基因完整编码区     cDNA序列,长度669 bp,编码222个氨基酸。经与谷胱甘肽转移酶家族其它成员的氨基酸序列比对和系统进化分析,初步确定PsGSTU1属于谷胱甘肽转移酶Tau家族。利用荧光定量RT-PCR分析表明,PsGSTU1在西伯利亚蓼的叶、茎和地下茎中均有表达,茎中表达量最高,叶和地下茎次之; 3% NaHCO3胁迫后,此基因在叶和茎中的最大表达量出现在胁迫后48 h,而茎在72 h;去胁迫后,此基因在不同部位的表达量表现为:地下茎>叶>茎。将PsGSTU1完整的cDNA片段连接至pET28a质粒以构建原核表达载体,并导入大肠杆菌(Escherichia coli),经IPTG诱导后表达了分子量为26 kD的目的蛋白,表达PsGSTU1的大肠杆菌显著提高了耐受NaHCO3胁迫的能力。
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刘明坤
刘关君
阎秀峰
刘昌财
魏志刚
刘桂丰
杨传平
关键词 西伯利亚蓼谷胱甘肽转移酶基因克隆RT-PCR表达分析    
AbstractGlutathione transferase (PsGSTU1) gene was isolated by RACE from Polygonum sibiricum Laxm, the PsGSTU1 gene had a complete open reading frame of 669 bp and encoded a peptide of 222 amino acid residues. Based on the comparison with other amino acids sequences among glutathione transferase family and the analysis of the phylogenetic evolution, this PsGSTU1 belonged to a glutathione transferase Tau kind of family. Expression analysis by RT-PCR showed that PsGSTU1 gene expressed in leaf, stem and root, especially higher expression in stems, next was leaf and root. After 3% NaHCO3 stress, this PsGSTU1 gene’s largest expression amount appeared at 48 h in leaf and root, but the largest expression within stem appeared at 72 h, When the stress was removed, the PsGSTU1 gene’s expression amount change within various organs were found as follows: root>leaf>stem. Subsequently, PsGSTU1 gene that had a complete cDNA was inserted into the pET28a plasmid to construct the prokaryotic vector and then transformed into Escherichia coli. The 26 kD protein was expressed  after inducing by IPTG, and the resistance of E. coli expressed protein to NaHCO3 got enhanced remarkably.
Key wordsPolygonum sibiricum Laxm.    glutathione transferase    gene clone    RT-PCR    expression analysis
收稿日期: 2008-05-07     
通讯作者: 刘关君   
引用本文:   
刘明坤;刘关君;阎秀峰;刘昌财;魏志刚;刘桂丰;杨传平. 西伯利亚蓼Tau类谷胱甘肽转移酶基因的克隆与表达分析[J]. , 2009, 17(1): 98-106.
链接本文:  
http://journal05.magtech.org.cn/Jwk_ny/CN/      或     http://journal05.magtech.org.cn/Jwk_ny/CN/Y2009/V17/I1/98
 
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