外源质粒电击转入Pseudomonas putida的条件研究  

Study on Electrotransformation of vitro DNA into Pseudomonas putida

作　　者：邓婷婷[1] 李晖[1] 刘燕[2] 郑元元[2] 李春[1,2] 

机构地区：[1]石河子大学农学院/新疆兵团化工绿色过程重点实验室,石河子832003 [2]北京理工大学生命科学与技术学院生物工程系,北京100081

出　　处：《中国生物工程杂志》2009年第8期81-85,共5页China Biotechnology

基　　金：国家自然科学基金(20776017);新疆维吾尔自治区高新技术重点项目(200811108)资助项目

摘　　要：以自行筛选的恶臭假单胞菌(Pseudomonas putida)(命名为Rs-198,Genbank登录号为FJ788425)为受体菌,将具有卡那霉素抗性标记的大肠杆菌-假单胞菌穿梭质粒PDSK519通过电转化法导入到受体菌中,对细胞生长状态、电转化温度、质粒DNA及感受态细胞浓度、电击电压及电转化介质给予转化效率的影响进行研究。结果表明,在细胞生长至OD600为0.5左右时收集菌体,在低温条件下制备浓度为4.6×1012/ml的感受态细胞,以0.3mol/L的蔗糖为电转化介质,在13kV/cm的场强下电击能获得较高的转化效率,最高可达1.3×107个转化子/μg DNA。为构建恶臭假单胞的遗传转化系统,利用基因工程手段为该菌的进一步研究奠定了理论基础。Pseudomonas putida isolated relieving salt stress and promoting plant growth was used as the recipient strain. Optimum conditions including growth stage of the strain, concentration of competent cell, electroshock voltage and the meduim were investigated for the electroporation of P. putida-E, coli shuher vector PDSKS19 into P. putida Rs-198. It was showed that the highest transformation efficiency was up to 1.3 × 10^7CFU/μL g DNA in the optimium condition, under which the competent cells were collected at logarithmic growth phase（ OD600 =0.5）, the mixture of the competent cells（4.6 ×10^12/ml） and PDSKS19 plasmid DNA was electroporated at 13kV/cm. It will be very helpful for developing a genetic transformation system and studying the genomic function of P. putida.

关 键 词：恶臭假单胞菌 电转化 转化效率 转化条件 外源质粒 

分 类 号：Q785[生物学—分子生物学]