萝卜水通道蛋白基因RsAQP1b的表达模式及功能分析

摘要

以萝卜品种白玉春为材料,利用同源克隆的方法获得萝卜水通道蛋白基因RsAQP1b的编码序列全长,在萝卜苗期干旱胁迫下分析RsAQP1b基因及蛋白的表达模式;在烟草中表达该基因,研究干旱胁迫下烟草中MDA含量及保护酶的活性变化.结果表明:萝卜水通道蛋白基因RsAQP1b的编码序列全长861 bp,编码286个氨基酸.制备了RsAQP1b多克隆抗体,效价在1:512000以上.Real time PCR与Western blot检测表明,干旱胁迫导致RsAQP16基因与蛋白的表达水平整体下降.PEG6000干旱处理下的转基因与野生型烟草MDA含量都增加,但是转基因烟草的增加幅度小;随着干旱程度加重,转mAQP1b基因烟草中SOD含量呈上升趋势,CAT含量呈先上升后下降趋势,POD含量呈上升趋势,酶活性均高于相应野生型对照.%Taking the radish cultivar ‘Baiyuchun’ as material,this paper gained coded sequence of aquaporin gene RsAQP1b by homology-based cloning method;and analyzed the expression pattern of RsAQP1b gene and protein under drought stress during radish seedling period,and studied the expression of this gene in tobacco,changes in MDA content and activity of protective enzyme under drought stress.The results indicated that CDS of RsAQP1b was 861 bp,encoding a protein of 286 amino acids.Polyclonal antibody of RsAQP1b was prepared and its titer was over 1 ∶ 512 000.Real time PCR and western blot detection showed that the expression level of RsAQP1b gene and protein was down-regulated by drought stress.Under PEG6000 drought stress treatment,the MDA content in transgenic and wild type (WT) tobacco was all increased,but the increasing rage of transgenic tobacco was small.The content of SOD in transgenic RsAQP1b tobacco showed an increasing tendency,while the CAT content showed a tendency of first rising then declining.The POD content showed an increasing tendency.The enzymatic activity was all exceeded that of the relevant CK of wide type.