Isotachophoresis for rapid transformation of Escherichia coli. Issue 4 (21st December 2021)

• Record Type: Journal Article
• Authors: Alves, Monica N.; Nai, Yi Heng; Powell, Shane M.; Macka, Mirek; Breadmore, Michael C.
• Is Part Of: Electrophoresis. Volume 43:Issue 4(2022)
• Page Start: 543
• Page End: 547
• Publication Date: 2021-12-21
• Subjects: Bacteria -- CE -- Isotachophoresis -- Plasmid -- Transformation; Electrophoresis -- Periodicals; Electrophoresis -- Periodicals; 541.372
• DOI: 10.1002/elps.202100095
• Languages: English
• ISSNs: 0173-0835

Abstract

Abstract: A frequent limitation of electroporation (EP) and chemical transformation (CT) are the need of tedious and time‐consuming procedures for inducing transformation competence, the substantial number of cells required, and the low transformation yields typically achieved. Here, we show a new and rapid electrokinetic method for transformation of small number of noncompetent Escherichia coli TOP10 cells (2–3 × 10 5 ) at room temperature. Escherichia coli TOP10 cells and plasmid DNA are sequentially injected into a 50 μm ID capillary and focused into 11.5 nL by isotachophoresis (ITP) induced by application of high DC voltage (–16 kV). Through ITP, a large excess of plasmid DNA is brought in contact with the cell surface, with the contact time adjusted by application of a counter‐pressure (1.3 psi) opposing the ITP movement. The transformation rate was more than 1000‐fold higher compared to EP and CT at survival rates greater than 60%. Abstract : Schematic of the ITP‐based transformation method showing transformed colonies obtained in absence and/or presence of counter‐pressure that opposes the voltage‐induced ITP movement.