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利用微冲击波进行细菌转化。

Bacterial transformation using micro-shock waves.

机构信息

Department of Aerospace Engineering, Indian Institute of Science, Bangalore 560 012, India.

出版信息

Anal Biochem. 2011 Dec 15;419(2):292-301. doi: 10.1016/j.ab.2011.08.038. Epub 2011 Aug 27.

Abstract

Shock waves are one of the most competent mechanisms of energy dissipation observed in nature. We have developed a novel device to generate controlled micro-shock waves using an explosive-coated polymer tube. In this study, we harnessed these controlled micro-shock waves to develop a unique bacterial transformation method. The conditions were optimized for the maximum transformation efficiency in Escherichia coli. The maximum transformation efficiency was obtained when we used a 30 cm length polymer tube, 100 μm thick metal foil, 200 mM CaCl(2), 1 ng/μl plasmid DNA concentration, and 1×10(9) cell density. The highest transformation efficiency achieved (1×10(-5) transformants/cell) was at least 10 times greater than the previously reported ultrasound-mediated transformation (1×10(-6) transformants/cell). This method was also successfully employed for the efficient and reproducible transformation of Pseudomonas aeruginosa and Salmonella typhimurium. This novel method of transformation was shown to be as efficient as electroporation with the added advantage of better recovery of cells, reduced cost (40 times cheaper than a commercial electroporator), and growth phase independent transformation.

摘要

冲击波是自然界中观察到的最有效的能量耗散机制之一。我们开发了一种使用爆炸涂层聚合物管产生受控微冲击波的新型装置。在这项研究中,我们利用这些受控的微冲击波开发了一种独特的细菌转化方法。优化了条件,以获得大肠杆菌的最大转化效率。当我们使用 30cm 长的聚合物管、100μm 厚的金属箔、200mM CaCl2、1ng/μl 质粒 DNA 浓度和 1×10(9)细胞密度时,获得了最大的转化效率。与之前报道的超声介导转化(1×10(-6)转化体/细胞)相比,获得的最高转化效率(1×10(-5)转化体/细胞)至少提高了 10 倍。该方法还成功地用于铜绿假单胞菌和鼠伤寒沙门氏菌的高效和可重复转化。与电穿孔相比,这种新的转化方法具有更好的细胞回收率、更低的成本(比商业电穿孔器便宜 40 倍)和生长阶段独立转化等优点。

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