苎麻酵母cDNA文库的构建与分析  

Construction and Analysis of cDNA Library of Ramie Yeast

作　　者：刘婕仪 苏小慧 全芮萍 王昕慧 刘皖慧 崔国贤[1] 杨瑞芳[1] 佘玮[1] Liu Jieyi;Su Xiaohui;Quan Ruiping;Wang Xinhui;Liu Wanhui;Cui Guoxian;Yang Ruifang;She Wei(College of Agronomy,Hunan Agricultural University,Changsha,410128)

机构地区：[1]湖南农业大学农学院,长沙410128

出　　处：《分子植物育种》2023年第13期4295-4300,共6页Molecular Plant Breeding

基　　金：国家自然科学基金项目(31871673);国家麻类产业技术体系养分管理岗位项目(CARS-16-E11)共同资助。

摘　　要：酵母cDNA文库构建是进行酵母双杂交的重要前提。获得高容量的苎麻(Boehmeria nivea L.)基因文库,能为深入挖掘苎麻重要性状基因、分子改良苎麻产量和品质提供坚实基础。本研究通过两种纯化方法构建了苎麻c DNA文库,利用SMART合成技术成功构建了高容量的苎麻酵母杂交cDNA文库。实验结果表明分段切胶回收纯化法的重组率和文库滴度分别为92%、2.5×107cfu/m L;SPIN+TE-400纯化双链cDNA纯化法得到的重组率和文库滴度分别为64%、1.32×10~7cfu/mL。分段切胶回收纯化双链cDNA在去除小片段、文库滴度、转化效率均SPIN+TE-400纯化双链cDNA纯化法好。两种方法构建的文库质量均达到酵母筛选文库标准,可用于苎麻基因表达调控的研究和互作蛋白的筛选试验。Construction of yeast cDNA library is an important prerequisite for yeast two-hybrid system.Obtaining high-capacity ramie(Boehmeria nivea L.)gene library can provide a solid foundation for further research of important trait genes and molecular improvement of yield and quality of ramie.The cDNA library of ramie was constructed by two purification methods,and the high-capacity cDNA library of ramie yeast hybrid was successfully constructed by SMART synthesis technique.The results showed that the recombinant rate and the titer of the library were 92%and 2.5×10^(7) cfu/mL,respectively.The recombinant rate and the titer of the library were 64%and 1.32×10^(7) cfu/mL.The double-stranded cDNA was purified by using the method of Spin+TE-400 with the removal of small fragments,library titer and transformation efficiency.The quality of the library constructed by the two methods was up to the standard of yeast screening library,which can be used for the study of regulation of gene expression and the screening test of interacting proteins.

关 键 词：苎麻(Boehmeria nivea L.) CDNA文库 SMART技术 

分 类 号：S563.1[农业科学—作物学]