少量制备大肠杆菌感受态细胞条件探索

目的:为了获得重复性好、转化率高的少量制备感受态细胞的方法,利用不同生长时期的大肠杆菌感受态细胞,进行转化比较。方法:根据普通实验室的实验条件,常规方法提取质粒,氯化钙法转化不同生长时期的大肠杆菌感受态细胞,比较转化率。结果:大肠杆菌感受态细胞的转化率与OD值显著相关,在OD600nm为0.39和0.55时转化率最高,在OD600nm为0.28~0.55之间均可得到理想的转化效果。结论:少量制备感受态细胞方法操作中无需添加任何保护试剂和细胞复苏培养,操作简便、重复性好,实验成本低廉。

Study on the Preparation of the Competent Cells of Escherichia coli

Objective: In order to get a method of competent cells preparation with well repeatability and high transformation efficiency,different growth phase of Escherichia coli was used to transform and compared their transformation efficiency.Methods: Draw the growth curve of Escherichia coli,choose the correct time to make competent cells,plasmid DNA was extracted using alkaline lysis method,transformed plasmid DNA into host cells using CaCl_2 method,compared the transformation ratio of different growth phase of Escherichia coli. Results: The transformation efficiency of E.coli was the highest in 0.39 and 0.55 of the OD_(600nm),and it showed that the competent cells could have better transformation efficiency when Escherichia coli grew up OD_(600nm) to 0.28-0.55.Conclusion: Because the preparation of the competent cells needed not any chemical reagents and anabiosis culture,the method of preparation of the competent cells of Escherichia coli was simple,repeatable and economical.