干酪乳杆菌LC2W最优电转化条件的建立

Establishment of genetic transformation system for Lactobacillus casei LC2W

中文关键词:  干酪乳杆菌  电转化  遗传转化  细胞壁弱化剂  电场强度

英文关键词:Lactobacillus casei  electransformation  genetic transformation  cell wall weakening agent  strength of electric field

中文摘要:

      目的 建立干酪乳杆菌LC2W电转化的最优条件。方法 研究了不同质粒、细胞壁弱化剂浓度、电场强度、复苏时间以及质粒浓度对电转效率的影响, 通过单因素轮换法进行了逐一优化。结果 经过对乳杆菌感受态制备及电转过程中各因素的考察, 得到最佳的参数条件为: 添加1 %甘氨酸作为细胞壁弱化剂制成感受态细胞, 电场强度为10 kV/cm进行高压电击后, 复苏3 h, 质粒浓度为200 ng/μL时, 能够达到最高的电转效率为1.25×106 CFU/μg DNA。结论 经优化后, 获得了较高的转化水平, 可用于指导多种乳杆菌的高效电转, 并且为下一步的LC2W菌株改造和基因功能探究奠定了基础。

英文摘要:

      Objective To establish a genetic transformation system of Lactobacillus casei LC2W. Methods On the basis of single factor experiments, the effects of operating conditions, such as different plasmids, concentration of cell wall weakening agent, strength of electric field, recovery time, and plasmid con-centration were investigated. Results The optimized parameters for electransformation were as follows: concentration of glycine used as cell wall weakening agent 1%, strength of electric field 10 kV/cm, recovery time 3 h, plasmid concentration 200 ng/μL. The high-efficient electransformation was reached at 1.25×106 CFU/μg DNA. Conclusions A high-efficient electransformation system was estabolished, which could be used as a protocol for other Lactobacilli and the basis for genetic modification of L. casei LC2W.