Abstract
We present a novel method for shockwave-assisted bacterial transformation using a miniature oxyhydrogen detonation-driven shock tube. We have obtained transformation efficiencies of about 1.28 × 106, 1.7 × 106, 5 × 106, 1 × 105, 1 × 105 and 2 × 105 transformants/µg of DNA for Escherichia coli, Salmonella Typhimurum, Pseudomonas aeruginosa, Mycobacterium smegmatis, Mycobacterium tuberculosis (Mtb) and Helicobacter pylori respectively using this method which are significantly higher than those obtained using conventional methods. Mtb is the most difficult bacteria to be transformed and hence their genetic modification is hampered due to their poor transformation efficiency. Experimental results show that longer steady time duration of the shockwave results in higher transformation efficiencies. Measurements of Young’s modulus and rigidity of cell wall give a good understanding of the transformation mechanism and these results have been validated computationally. We describe the development of a novel shockwave device for efficient bacterial transformation in complex bacteria along with experimental evidence for understanding the transformation mechanism.
Highlights
Transferring DNA/RNA to the cells is an important requisite for basic as well as applied biotechnology and molecular biology studies
High transformation efficiencies were obtained in bacterial strains of Escherichia coli, Salmonella Typhimurium, Pseudomonas aeruginosa, Mycobacterium smegmatis, Mycobacterium tuberculosis and Helicobacter pylori
We report the development of a novel shockwave generator, its application for bacterial transformation and the mechanical changes caused by shockwave exposure and its importance in the mechanism of efficient transformation
Summary
Transferring DNA/RNA to the cells is an important requisite for basic as well as applied biotechnology and molecular biology studies. Many dry particle delivery systems have been developed using high pressure gases to accelerate particles for plant transformation[5, 6] Shockwaves prove their importance and role in inducing cell competence. A novel shock wave-induced transfer of DNA into bacteria using dual-pulse (tandem) shockwaves with 50-fold higher transformation efficiency by enhancing cavitation is reported[9]. It was shown that the impulse of the shock wave (i.e., the pressure integrated over time) plays a more dominant role in creating cell permeability rather than the peak pressure of the shockwave This device demonstrates potential for delivery of exogenous DNA through cell wall of simple as well as complex bacterial strains. We report the development of a novel shockwave generator, its application for bacterial transformation and the mechanical changes caused by shockwave exposure and its importance in the mechanism of efficient transformation
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