摘要： 将带有目的基因SeNHX1、GFP的酵母表达载体pYES2-SeNHX1、pYES2-GFP、pYES2-SeNHX1-GFP分别导入nhx1酵母缺失型菌株296H,用共聚焦荧光显微镜观察SeNHX1表达蛋白的亚细胞定位;通过菌株在含盐培养基中生长状况研究该蛋白对Na+的解毒功能。结果表明,296H(pYES2-GFP)菌株的绿色荧光弥散于整个细胞质,而296H(pYES2-SeNHX1-GFP)菌株的绿色荧光包围于液泡膜的一圈,因此,SeNHX1蛋白定位于液泡膜;296H(pYES2-GFP)和296H缺陷型菌株在含0.5 mol/L NaCl的培养基中生长受到抑制;带有SeNHX1目的基因的转化子比以上2种菌株在含盐培养基中生长更快,表明SeNHX1基因可恢复nhx1酵母突变体Na+的解毒功能。SeNHX1疏水性分析、氨基酸序列分析、亚细胞定位及其对Na+的解毒功能结果说明,SeNHX1基因是可提高生物体耐盐性的液泡膜Na+/H+逆向转运蛋白基因。
关键词:
SeNHX1基因,
酵母,
表达载体,
GFP绿色荧光,
耐盐性,
亚细胞定位
Abstract: Yeast expression vector pYES2-SeNHX1,pYES2-GFP,pYES2-SeNHX1-GFP were introduced into the nhx1 mutant strain 296H.The protein localization of SeNHX1 was observed by confocal laser scanning microscope and the function of Na＋-detoxification was studied by growth of 296H with different expression vector in medium containing 0.5 mol/L NaCl.The results showed that green fluorescence dispersed in the whole cell in 296H(pYES2-GFP),but only appeared on the vacuolar membrane and shaped a cyclic structure in 296H(pYES2-SeNHX1-GFP).So SeNHX1 was guessed to be localized on the vacuolar membrane.The strains of 296H(pYES2-GFP) and 296H were inhibited for growth when cultured in medium with NaCl,where the strains with SeNHX1 gene grew better.The finding suggested SeNHX1 gene could complement the function of nhx1 mutant.Therefore,on the base of amino acid sequence analysis and hydrophobicity analysis,SeNHX1 was confirmed as vacuolar Na＋ /H＋ antiporters,which conferred to the salt tolerance of plants.
Key words:
SeNHX1 gene,
Yeast,
Expression vector,
GFP green fluorescence,
Salt-tolerance,
SeNHX1 localization.
中图分类号:
杨小玲, 季静, 王罡. SeNHX1表达蛋白的亚细胞定位及其对Na~+的解毒功能[J]. 华北农学报, 2012, 27(5): 8-11. doi: 10.3969/j.issn.1000-7091.2012.05.002.
YANG Xiao-ling, JI Jing, WANG Gang. SeNHX1 Protein Localization and Its Function of Na~＋-detoxification[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2012, 27(5): 8-11. doi: 10.3969/j.issn.1000-7091.2012.05.002.