Ultrasound-mediated DNA transfer for bacteria

Y. Song, T. Hahn, I. P. Thompson, Timothy J. Mason, G. M. Preston, G. Li, Larysa Paniwnyk, W. E. Huang

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Abstract

In environmental microbiology, the most commonly used methods of bacterial DNA transfer are conjugation and electroporation. However, conjugation requires physical contact and cell–pilus–cell interactions; electroporation requires low-ionic strength medium and high voltage. These limitations have hampered broad applications of bacterial DNA delivery. We have employed a standard low frequency 40 kHz ultrasound bath to successfully transfer plasmid pBBR1MCS2 into Pseudomonas putida UWC1, Escherichia coli DH5α and Pseudomonas fluorescens SBW25 with high efficiency. Under optimal conditions: ultrasound exposure time of 10 s, 50 mM CaCl2, temperature of 22°C, plasmid concentration of 0.8 ng/µl, P. putida UWC1 cell concentration of 2.5 × 109 CFU (colony forming unit)/ml and reaction volume of 500 µl, the efficiency of ultrasound DNA delivery (UDD) was 9.8 ± 2.3 × 10−6 transformants per cell, which was nine times more efficient than conjugation, and even four times greater than electroporation. We have also transferred pBBR1MCS2 into E. coli DH5α and P. fluorescens SBW25 with efficiencies of 1.16 ± 0.13 × 10−6 and 4.33 ± 0.78 × 10−6 transformants per cell, respectively. Low frequency UDD can be readily scaled up, allowing for the application of UDD not only in laboratory conditions but also on an industrial scale.
Original languageEnglish
Article numbere129
JournalNucleic Acids Research
Volume35
Issue number19
DOIs
Publication statusPublished - 22 Sep 2007

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Electroporation
Bacterial DNA
Pseudomonas fluorescens
Pseudomonas putida
Bacteria
DNA
Plasmids
Environmental Microbiology
Escherichia coli
Baths
Osmolar Concentration
Stem Cells
Temperature

Bibliographical note

The full text is also available from: http://nar.oxfordjournals.org/content/35/19/e129.full.pdf+html
This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/2.0/uk/) which permits unrestricted non-commercial use, distribution, and reproduction in any medium, provided the original work is properly cited.

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Song, Y., Hahn, T., Thompson, I. P., Mason, T. J., Preston, G. M., Li, G., ... Huang, W. E. (2007). Ultrasound-mediated DNA transfer for bacteria. Nucleic Acids Research, 35(19), [e129]. https://doi.org/10.1093/nar/gkm710

Ultrasound-mediated DNA transfer for bacteria. / Song, Y.; Hahn, T.; Thompson, I. P.; Mason, Timothy J.; Preston, G. M.; Li, G.; Paniwnyk, Larysa; Huang, W. E.

In: Nucleic Acids Research, Vol. 35, No. 19, e129, 22.09.2007.

Research output: Contribution to journalArticle

Song, Y, Hahn, T, Thompson, IP, Mason, TJ, Preston, GM, Li, G, Paniwnyk, L & Huang, WE 2007, 'Ultrasound-mediated DNA transfer for bacteria' Nucleic Acids Research, vol. 35, no. 19, e129. https://doi.org/10.1093/nar/gkm710
Song Y, Hahn T, Thompson IP, Mason TJ, Preston GM, Li G et al. Ultrasound-mediated DNA transfer for bacteria. Nucleic Acids Research. 2007 Sep 22;35(19). e129. https://doi.org/10.1093/nar/gkm710
Song, Y. ; Hahn, T. ; Thompson, I. P. ; Mason, Timothy J. ; Preston, G. M. ; Li, G. ; Paniwnyk, Larysa ; Huang, W. E. / Ultrasound-mediated DNA transfer for bacteria. In: Nucleic Acids Research. 2007 ; Vol. 35, No. 19.
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N2 - In environmental microbiology, the most commonly used methods of bacterial DNA transfer are conjugation and electroporation. However, conjugation requires physical contact and cell–pilus–cell interactions; electroporation requires low-ionic strength medium and high voltage. These limitations have hampered broad applications of bacterial DNA delivery. We have employed a standard low frequency 40 kHz ultrasound bath to successfully transfer plasmid pBBR1MCS2 into Pseudomonas putida UWC1, Escherichia coli DH5α and Pseudomonas fluorescens SBW25 with high efficiency. Under optimal conditions: ultrasound exposure time of 10 s, 50 mM CaCl2, temperature of 22°C, plasmid concentration of 0.8 ng/µl, P. putida UWC1 cell concentration of 2.5 × 109 CFU (colony forming unit)/ml and reaction volume of 500 µl, the efficiency of ultrasound DNA delivery (UDD) was 9.8 ± 2.3 × 10−6 transformants per cell, which was nine times more efficient than conjugation, and even four times greater than electroporation. We have also transferred pBBR1MCS2 into E. coli DH5α and P. fluorescens SBW25 with efficiencies of 1.16 ± 0.13 × 10−6 and 4.33 ± 0.78 × 10−6 transformants per cell, respectively. Low frequency UDD can be readily scaled up, allowing for the application of UDD not only in laboratory conditions but also on an industrial scale.

AB - In environmental microbiology, the most commonly used methods of bacterial DNA transfer are conjugation and electroporation. However, conjugation requires physical contact and cell–pilus–cell interactions; electroporation requires low-ionic strength medium and high voltage. These limitations have hampered broad applications of bacterial DNA delivery. We have employed a standard low frequency 40 kHz ultrasound bath to successfully transfer plasmid pBBR1MCS2 into Pseudomonas putida UWC1, Escherichia coli DH5α and Pseudomonas fluorescens SBW25 with high efficiency. Under optimal conditions: ultrasound exposure time of 10 s, 50 mM CaCl2, temperature of 22°C, plasmid concentration of 0.8 ng/µl, P. putida UWC1 cell concentration of 2.5 × 109 CFU (colony forming unit)/ml and reaction volume of 500 µl, the efficiency of ultrasound DNA delivery (UDD) was 9.8 ± 2.3 × 10−6 transformants per cell, which was nine times more efficient than conjugation, and even four times greater than electroporation. We have also transferred pBBR1MCS2 into E. coli DH5α and P. fluorescens SBW25 with efficiencies of 1.16 ± 0.13 × 10−6 and 4.33 ± 0.78 × 10−6 transformants per cell, respectively. Low frequency UDD can be readily scaled up, allowing for the application of UDD not only in laboratory conditions but also on an industrial scale.

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