黄瓜矮化突变体膨胀素Cs-EXPA1蛋白的表达及活性分析  

Expression of the dwarf cucumis mutant expansin Cs-EXPA1 protein and analysis of its activity

作　　者：胡宝忠[1] 黄莎莎[1] 李凤兰[1] 王多佳[1] 

机构地区：[1]东北农业大学生命科学学院,哈尔滨150030

出　　处：《东北农业大学学报》2013年第10期44-50,共7页Journal of Northeast Agricultural University

基　　金：黑龙江省自然科学基金项目(230340);高等学校博士点专项科研基金(200802240008)

摘　　要：摘要：采用RT—PCR法从矮化黄瓜D0462中分离膨胀素基因Cs—EXPAl，并将其与毕赤酵母分泌型表达载体pGAPHoLM连接，形成重组质粒pGAPHcrM-EXPAl。通过电击转化法将重组质粒转化到巴氏德毕赤酵母GS115中，经检测膨胀素基因通过同源重组整合到毕赤酵母染色体上，成功获得重组菌株。该茵株在摇床培养条件下可在上清液中检测到膨胀素蛋白。该蛋白不具有单独水解纤维素的活性，但在纤维素酶水解纤维素时却具有协同水解纤维素的活性。文章初步确定膨胀素协同水解纤维素的最佳时间36h、最适pH4．0．5．0、最适温度50qC。研究可为提高纤维素的酶水解效率提供理论参考。The cDNA of expansin Cs-EXPA1 was isolated from Dwarf Cucumis D0462 through RT-PCIR, it was ligated with the Pichia secretion expression vector pGAPHa, and resulting in the recombinant plasmid pGAPHaM-EXPA1. The recombinant plasmid was transformed into P. pastoris GSl15 through electroporation. The expansin gene was in frame integrated into the Pichia genome through homologous recombination and resulted recombinant strain. With shake cultivation, we can detect expansin protein in supernatant. This protein don＇t have activity of hydrolysis cellulose alone, but it have synergistic activity of hydrolysis cellulose when cellulase hydrolysis cellulose. We definited optimization time, tempreture and pH of synergistic hydrolysis cellulose through our experiment. Our study willraise hydrolysis efficiency and to establish foundation for research high efficiency hydrolysis process of cellulase.

关 键 词：膨胀素 毕赤酵母 黄瓜 纤维素 协同作用 

分 类 号：TS255.3[轻工技术与工程—农产品加工及贮藏工程]