西伯利亚蓼谷胱甘肽转移酶和半胱氨酸合成酶基因在酿酒酵母中的共表达

谷胱甘肽转移酶和半胱氨酸合成酶在清除活性氧(reactive oxygen species,ROS)中起重要作用。采用0.36 mol.L-1NaHCO3对西伯利亚蓼(Polygonum sibiricum)进行胁迫处理, 荧光定量PCR分析表明这2个基因的表达受盐胁迫强烈诱导。为了分析2个基因是否具有抗盐能力以及其相互协同能力, 从cDNA文库中获得谷胱甘肽转移酶(GST)和半胱氨酸合成酶(CS)2个基因, 分别将GST、CS和GST+CS转入酿酒酵母(Saccharomyces cerevisiae)中, 并分别命名转基因酵母为ty-gst、tycs和ty-gc。在1 mol.L-1 Na2CO3和5 mol.L-1 NaCl胁迫处理下, 转基因酵母(ty-gst、ty-cs和ty-gc)的耐盐能力均明显高于野生型酵母(wy), 而三者之间并无显著差别。在0.4 mol.L-1 NaCl胁迫处理下, 转基因酵母(ty-gst、ty-cs和ty-gc)的抗氧化酶类相关基因SOD1、SOD2、GPX1和GPX3的表达量均低于野生型酵母(对照)(wy), 而CTA1表达量均高于野生型酵母(对照)(wy)。转基因酵母ty-cs在0.4 mol.L-1 NaCl胁迫处理前后其超氧化物歧化酶(superoxide dismutase, SOD)、过氧化氢酶(catalase, CAT)和谷胱甘肽过氧化物酶(glutathione peroxidase, GPX)的活性均表现为最高。

Co-expression of Polygonum sibiricum Glutathione Transferase and Cysteine Synthase Genes in Saccharomyces cerevisiae

Glutathione transferase (GST) and cysteine synthase (CS) play an important role in plant elimination of active oxygen (reactive oxygen species, ROS). RT-PCR analysis of GST and CS in Polygonum sibiricum Laxm. showed remarkable change in expression induced by 3% NaHCO3 stress. We transformed GST, CS and the GST+CS combination into yeast cells(Saccharomyces cerevisiae), designated transgenic yeast (ty) ty-gst, ty-cs and ty-gc. The survival rate of ty-gst, ty-cs and ty-gc was higher than that of wild-type yeast (wy) under 1 mol.L-1 Na2CO3 and 5 mol.L-1 NaCl stress, with essentially no difference in rate between the three tys. With 0.4 mol.L-1 NaCl, the activity of superoxide dismutase 1 (SOD1), SOD2, glutathione peroxidase 1 (GPX1) and GPX3 were lower and that of catalase 1 (CTA1) higher in ty than in wy. However, the activity of the ty-cs SOD, CAT and GPX showed the highest with 0.4 mol.L-1 NaCl stress.