菠菜甜菜碱醛脱氢酶因在棉花中的过量表达和抗冻耐逆性分析.docVIP

菠菜甜菜碱醛脱氢酶基因在棉花中的过量表达和抗冻耐逆性分析 罗晓丽1, 肖娟丽1, 王志安1, 张安红1, 田颖川2, 吴家和1,2 1 山西省农业科学院棉花研究所, 运城 0440002 中国科学院微生物研究所植物基因组国家重点实验室, 北京 100101 摘 要: 分离出菠菜甜菜碱醛脱氢酶基因(SoBADH)构建成由CaMV35S驱动的双元植物表达载体pBSB, 农杆菌菌株LBA4404携带该载体转化棉花, 获得转基因棉花植株。65株转基因植株经过PCR筛选、Southern blotting分析证明有45株为成功的转化株, 外源基因已经被整合到棉花的染色体组中, 并以单拷贝插入居多。对部分株系的SoBADH基因的表达进行分析表明均有较高的mRNA和蛋白的表达。经测定这些株系中的甜菜碱脱氢酶活性显著提高, 达到0.66~ 1.70 nmol/min/mg水平。同时这些转基因株系在盐胁迫下比对照长势强, 株高和地上部分的鲜重显著高于非转基因对照; 在低温胁迫下, 这些转基因株系表现出显著的抗冻性能。结果表明菠菜甜菜碱醛脱氢酶能够在异源植物棉花中过量表达, 并具有较高的酶活性, 转基因棉花可作为抗逆育种的种质材料。 关键词: 棉花, 菠菜甜菜碱醛脱氢酶, 耐盐性, 转基因 Overexpression of Spinacia oleracea Betaine AldehydeDehydrogenase (SoBADH) Gene Confers the Salt and Cold Tolerant in Gossypium hirsutum L. Xiaoli Luo1, Juanli Xiao1, Zhian Wang1, Anhong Zhang1, Yingchuan Tian2, and Jiahe Wu1,2 1 Institute of Cotton Research, Shanxi Academy of Agricultural Science, Yuncheng 044000, China 2 State Key Laboratory of Plant Genomics, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China Abstract: The open reading frame of Spinacia oleracea Betaine Aldehyde Dehydrogenase (SoBADH) was retrieved from Spinacia oleracea and inserted into the Agrobacterium tumefaciens binary vector pBin438, which was driven by CaMV35S promoter, and produced the new binary vector pBSB. A. tumefaciens LBA4404 carrying this plasmid was used in genetic transformation of plants. Forty-five primary transgenic plants were detected by PCR and verified by the Southern blotting from 65 regenerated plants, of which 27 transgenic plants had only one copy of T-DNA. The Northern blotting and Western blotting analysis indicated that the SoBADH gene had been transcribed mRNA and expression protein in the transgenic cotton lines. The testing of SoBADH activity of transgenic plant leaves showed that the enzyme activity was much higher than that of the non-transgenic cotton. The growth of transgenic plants was well under the salinity and freezing stress,