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Isotachophoresis for rapid transformation of Escherichia coli
journal contributionposted on 2023-05-21, 15:30 authored by Alves, MN, Nai, YN, Shane PowellShane Powell, Miroslav MackaMiroslav Macka, Michael BreadmoreMichael Breadmore
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 × 105 ) 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%.
Department/SchoolSchool of Natural Sciences
Place of publicationGermany