西伯利亚蓼谷氨酰胺合成酶基因的克隆及碱性盐胁迫下的表达

Cloning of Coding Sequence of Glutamin Synthetase from Polygonum sibiricum Laxm. and Its Expression under Alkali Salinity Stress

根据西伯利亚蓼(Polygonum sibiricum Laxm.)地下茎抑制消减文库(SSH)中获得的谷氨酰胺合成酶基因(Glutamin synthetase,GS)EST序列,应用RACE技术克隆了具有Poly A的全长cDNA序列,以下简称为PsGS基因。该序列全长1 273 bp,其5'非翻译区178 bp,3'非翻译区24 bp,开放阅读框编码356个氨基酸残基;根据与其他植物谷氨酰胺合成酶的氨基酸序列的比对以及系统进化分析的结果,确定此基因为谷氨酰胺合成酶基因家族成员;经过SignalP3.0预测该蛋白没有信号肽,无切割位点,为非分泌蛋白。经过ProtParam计算该蛋白的理论等电点为5.55,分子量为39.2 kD,不稳定系数为43.82%,为非稳定蛋白。实时定量PCR分析表明,PsGS在西伯利亚蓼叶、茎、地下茎中均有表达。在3%NaHCO3诱导下,该基因在叶和茎中表达升高,在地下茎中表达受到抑制,推测该基因在抵御碱性盐迫时具有重要作用。

The full-length of Glutamin synthetase （termed PsGS） gene was cloned using RACE （rapid amplification of cDNA end） technology based on GS partial sequence obtained from a random clone in SSH library of Polygonum sibiricum Laxm.. The acquired 1 273 bp sequence includes a 5′ untranslated region of 178 bp, a 3′ untranslated region of 24 bp with poly （A）, and an open reading frame （ORF） encoding 356 amino acids. Based on the comparison with amino acid sequences of other plant glutamin synthetase and the phylogenetic analysis of protein evolution, this gene was divided into glutamin synthetase family. The expression analyses by RT-PCR showed that PsGS expressed in leaves, stems and rhizomes of P.sibiricum Laxm.. Under the induction of 3% NaHCO3, the expression of PsGS was significantly influenced, which suggested that PsGS might play an important role in alkali salt stress resistance.