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Nucleic acid and protein extraction from electropermeabilized E. coli cells on a microfluidic chip.

Matos, Tiago LU ; Senkbeil, S; Mendonça, A; Queiroz, J A; Kutter, J P and Bülow, Leif LU (2013) In Analyst 138(24). p.7347-7353
Abstract
Due to the extensive use of nucleic acid and protein analysis of bacterial samples, there is a need for simple and rapid extraction protocols for both plasmid DNA and RNA molecules as well as reporter proteins like the green fluorescent protein (GFP). In this report, an electropermeability technique has been developed which is based on exposing E. coli cells to low voltages to allow extraction of nucleic acids and proteins. The flow-through electropermeability chip used consists of a microfluidic channel with integrated gold electrodes that promote cell envelope channel formation at low applied voltages. This will allow small biomolecules with diameters less than 30 A to rapidly diffuse from the permeabilized cells to the surrounding... (More)
Due to the extensive use of nucleic acid and protein analysis of bacterial samples, there is a need for simple and rapid extraction protocols for both plasmid DNA and RNA molecules as well as reporter proteins like the green fluorescent protein (GFP). In this report, an electropermeability technique has been developed which is based on exposing E. coli cells to low voltages to allow extraction of nucleic acids and proteins. The flow-through electropermeability chip used consists of a microfluidic channel with integrated gold electrodes that promote cell envelope channel formation at low applied voltages. This will allow small biomolecules with diameters less than 30 A to rapidly diffuse from the permeabilized cells to the surrounding solution. By controlling the applied voltage, partial and transient to complete cell opening can be obtained. By using DC voltages below 0.5 V, cell lysis can be avoided and the transiently formed pores can be closed again and the cells survive. This method has been used to extract RNA and GFP molecules under conditions of electropermeability. Plasmid DNA could be recovered when the applied voltage was increased to 2 V, thus causing complete cell lysis. (Less)
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author
organization
publishing date
type
Contribution to journal
publication status
published
subject
in
Analyst
volume
138
issue
24
pages
7347 - 7353
publisher
Royal Society of Chemistry
external identifiers
  • wos:000326988500012
  • pmid:24162237
  • scopus:84887544786
ISSN
1364-5528
DOI
10.1039/c3an01576a
language
English
LU publication?
yes
id
017b1436-c08d-4fd4-9083-2c90a4742372 (old id 4142974)
date added to LUP
2013-11-12 17:51:45
date last changed
2019-02-20 02:13:20
@article{017b1436-c08d-4fd4-9083-2c90a4742372,
  abstract     = {Due to the extensive use of nucleic acid and protein analysis of bacterial samples, there is a need for simple and rapid extraction protocols for both plasmid DNA and RNA molecules as well as reporter proteins like the green fluorescent protein (GFP). In this report, an electropermeability technique has been developed which is based on exposing E. coli cells to low voltages to allow extraction of nucleic acids and proteins. The flow-through electropermeability chip used consists of a microfluidic channel with integrated gold electrodes that promote cell envelope channel formation at low applied voltages. This will allow small biomolecules with diameters less than 30 A to rapidly diffuse from the permeabilized cells to the surrounding solution. By controlling the applied voltage, partial and transient to complete cell opening can be obtained. By using DC voltages below 0.5 V, cell lysis can be avoided and the transiently formed pores can be closed again and the cells survive. This method has been used to extract RNA and GFP molecules under conditions of electropermeability. Plasmid DNA could be recovered when the applied voltage was increased to 2 V, thus causing complete cell lysis.},
  author       = {Matos, Tiago and Senkbeil, S and Mendonça, A and Queiroz, J A and Kutter, J P and Bülow, Leif},
  issn         = {1364-5528},
  language     = {eng},
  number       = {24},
  pages        = {7347--7353},
  publisher    = {Royal Society of Chemistry},
  series       = {Analyst},
  title        = {Nucleic acid and protein extraction from electropermeabilized E. coli cells on a microfluidic chip.},
  url          = {http://dx.doi.org/10.1039/c3an01576a},
  volume       = {138},
  year         = {2013},
}