磁珠分离法快速提取大肠杆菌质粒DNA

Rapid Extraction of Plasmid DNA from Escherichia coli Using Magnetic Bead Separation Method

以单分散羧基功能化纳米磁性粒子为固相载体,PEG/NaCl为结合液,对大肠杆菌(Escherichia coli)裂解液中的质粒DNA进行了纯化研究。结果表明:PEG8000质量分数7.5%,NaCl浓度1.25 mol/L,磁珠用量0.9 mg时,在室温下,从1.5 mL大肠杆菌菌液中提取到的质粒DNA量为8.3μg,其A260/A280比值稳定在1.8左右,耗时20 min。与传统方法相比较,本提取法具有高效、便捷、经济和安全等优点,不仅适合于实验室的小量质粒DNA的提取,也适合于自动化DNA分析检测平台的构建。

In this paper,carboxyl-group functionalized single-dispersed magnetic nanoparticles were used as solid phase to extract plasmid DNA from crude Escherichia coli lysate by using PEG/NaCl as binding buffer.The results indicated that when the PEG/NaCl binding buffer was 7.5%/1.25 mol/L,at room temperature,8.3 μg of plasmid DNA which A260/A280 ratio stabilized at around 1.8 can be harvested from 1.5 mL overnight cultured E.coli bacterium by using 0.9 mg magnetic nanoparticles.The time needed was only 20 min.Compared with conventional methods,the magnetic bead separation method is efficient,convenient,economic and security,not only suitable for small scale preparation of pDNA at laboratory,but also can be adopted to construct automated pDNA analysis and identification platforms.