| 乳酸乳球菌电击转化方法的优化 |
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| 引用本文： | 刘亚欣,王宇,杨诺,郭桂英,李迁,曾纪锋,郑继平.乳酸乳球菌电击转化方法的优化[J].热带生物学报,2020,11(2):245-250. |
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| 作者姓名： | 刘亚欣 王宇 杨诺 郭桂英 李迁 曾纪锋 郑继平 |
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| 作者单位： | 1.海南大学 生命科学与药学院，海口 570228 |
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| 基金项目： | 国家自然科学基金（31460699）；海南自然科学基金创新研究团队项目（2017CXTD005）；海南自然科学基金面上项目（317071）；海南大学研究生教学经费（J100029019106004001） |
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| 摘 要： | 乳酸菌NICE系统是食品级表达系统，本实验以该系统的表达载体pNZ8148和宿主菌NZ9000为研究对象，对电击转化方法进行了优化。结果表明，在培养基中加入2.5%甘氨酸，收集OD600值为0.3的细胞、用配方I（洗涤液I:蔗糖0.5 mol·L?1+甘油10%；洗涤液II:蔗糖0.5 mol·L?1+甘油10%+EDTA50 mmol·L?1）]洗涤细胞，再转入0.2 μg质粒，利用2.5 kV电压和0.2 cm电击杯进行电击转化，恢复培养1.5 h后转化效率最高，达1.6×108 CFU·μg?1。本实验为乳酸乳球菌NZ9000的电击转化参数提供了数据参考，为基于乳酸菌NICE系统的深入研究奠定了基础。
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| 关 键 词： | 乳酸乳球菌 电击转化 电击转化优化 |
| 收稿时间： | 2019-12-07 |
Optimization of Electrotransformation of Lactococcus lactis |
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| Institution: | 1.Institute of Life Sciences and Pharmacy, Hainan University, Haikou, Hainan 5702282.Academic Affairs Office, Hainan University, Haikou, Hainan 5702283.Network and Technology Center, Hainan University, Haikou, Hainan 5702284.Institute of Animal Science and Technology, Hainan University, Haikou, Hainan 570228, China |
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| Abstract: | The nisin-controlled gene expression system (NICE) is a food-grade system used for gene expression in Lactococcus lactis. Electrotransformation was optimized via the NICE, consisting of an expression vector pNZ8148 and a host strain Lactococcus lactis NZ9000. The results showed that the electrotransformation of L. lactis NZ9000 with the expression vector pHZ8148 gave the highest efficiency (1.6 × 108 CFU μg?1) when L. lactis NZ9000 was cultured on the medium added with 2.5% of glycine, and its competent cells grown to 0.3 OD600 were prepared by washing with the liquid detergent formulation I (Sucrose 0.5 mol·L?1+glycerol 10%), transferred into with 0.2 μg plasmid of pNA8148, electrotranformed at 2.5 kV on a 0.2 cm gap cuvette, and mixed gently for 1.5 h before plating. This study provides optimum conditions for electrotransformation of L. lactis NZ9000 which are favorable for further study of L. lactis based on the NICE. |
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