电穿孔法介导质粒DNA转化大肠杆菌XL1-Blue MRF′菌株的实验研究

Experimental research on transformation of plasmid DNA into Escherichia coli XL1-Blue MRF′ strain mediated by electroporation.

目的探讨电穿孔法介导质粒DNA转化大肠杆菌XL1-Blue MRF′的最佳条件,提高其转化效率。方法将pCMV-Scrip质粒转化大肠杆菌XL1-Blue MRF′菌株,观察不同的电转化条件对转化效率的影响。结果电场强度、脉冲时间、电击缓冲液的离子强度、细菌生长状态、感受态细胞保存时间、质粒DNA浓度对转化效率均有不同程度的影响;在电压2.0kV,电阻200Ω,电容25μF,脉冲时间4.3ms和低离子强度电击缓冲液的条件下转化,能获得较高的转化率。结论优化电穿孔条件能提高转化效率。

Objective To explore the optimal protocol of transformation of plasrnid DNA into Escherichia coli（E, coli）XL1-Blue MRF＇ mediated by electroporation and improve its transformation efficiency. Methods Plasmid pCMV-Scrip was transferred into E. coli XL1-Blue MRF＇ strain by electroporation using different electrotransformation conditions and transformation efficiency was observed. Results The transformation efficiency was affected by electrical field strength,pulse time,ionic strength of electroshock buffer,growth phase of germ, storage duration of competent cells and concentration of plasmid DNA. The optimal transformation parameters for highest transformation efficiency were voltage 2.0kV, electric resistance 200Ω, capacitance 25μF, pulse time 4.3ms and electrotransformation buffer with low ionic strength. Conclusion Optimized electrotransformation conditions can improve transformation efficiency.