利用组成型表达载体pMG36e电转化乳酸乳球菌ML23的研究  

Research of electroporation processing of Lactococcus lactis ML23 with the constitutive expression vector pMG36e

作　　者：张玉兰[1] 柯晓静[2] 郭红敏[1] 谷新晰[1] 田洪涛[1] 郭兴华[3] 罗云波[4] 

机构地区：[1]河北农业大学食品科技学院,河北保定071001 [2]河北农业大学研究生学院,河北保定071001 [3]中国科学院微生物研究所,北京100080 [4]中国农业大学食品科技与营养工程学院,北京100083

出　　处：《河北农业大学学报》2010年第4期71-76,共6页Journal of Hebei Agricultural University

基　　金：国家高技术研究发展计划(863计划)项目(2006AA10Z317)

摘　　要：为了探明乳酸菌电转化的最适条件,提高电转化效率,本试验以大肠杆菌-乳酸菌穿梭型质粒pMG36e为载体,以乳酸乳球菌ML23为受体,利用电转化方法研究了受体菌株的生长状态与细胞壁处理方式、质粒浓度、电场强度、复苏培养基组成与复苏培养时间、受体菌株的限制修饰作用对电转化效率的影响。结果表明:含2%甘氨酸和0.5 mol/L蔗糖的MRS培养基培养至对数生长中期的细胞,在电场强度11.0 kV/cm、电阻200Ω、电容25μF下电击转化,转化后细胞在SMRS培养基中培养2 h,获得了较高的电转化效率。采用受体菌株修饰的质粒进行电转化,转化效率又提高了15倍以上,达1.05×105CFU/μg DNA。In order to improve the electroporation efficiency of lactic acid bacteria and optimize the conditions for electroporation of Lactococcus lactis ML23,a shuttle vector for Escherichia coli and Lactobacillus,pMG36e,were used.Factors including wall treatment,plasmid concentration,electric filed strength,composition of the recovery medium,recovery culture time and the restriction modification of host were investigated.Results showed that the optimal values for concentration of glycine and of sucrose and the pulse strength,risistance are 2.0%（W/V）,0.5 mol/L and 11 kV/cm,200 Ω,respectively.Best electroporation efficiency is attained after incubation in 0.5 mol/L sucrose containing MRS broth for another 2 h.When the plasmid DNA carrying the host strain modification was used to electroporate strain ML23,the transformation efficiency increased by 15 times,and reached 1.05×105 CFU /μg DNA.

关 键 词：乳酸乳球菌ML23 组成型表达载体 质粒pMG36e 电转化 电转化效率 

分 类 号：Q785[生物学—分子生物学]