title: Transformation of Escherichia coli JM109 using pUC19 by the Yoshida effect.
abstract: Transformation of non-competent Escherichia coli JM109 was accomplished using pUC19 as donor plasmid and sepiolite as the acicular material to promote cell piercing via application of friction with a polystyrene stick or a magnetic bar on the surface of a hydrogel containing agar. An automatic spreading setup was built with a conventional stirring plate and compared to manual spreading. Several parameters were optimized, namely, the agar content of the hydrogel (2%), concentration of cells (OD=1.3 corresponding to 1.4×10(9) bacterial cells/mL), concentration of sepiolite (0.01%), manual versus mechanical spreading (automatic spreading more consistent) and spreading time (30s). Efficiency values up to 4.1×10(4) CFU/μg pUC19 were obtained. The method proved to be suitable for a rapid and low cost transformation of non-competent E. coli JM109, where higher values of efficiency do not need to be attained.
authors.affiliation: CEB - Centro de Engenharia Biológica, Universidade do Minho, 4710-057 Braga, Portugal; Centro/Departamento de Física, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: G P
authors.initials: GP
authors.lastname: Mendes
authors.affiliation: CEB - Centro de Engenharia Biológica, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: P S
authors.initials: PS
authors.lastname: Vieira
authors.affiliation: Centro/Departamento de Física, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: S
authors.initials: S
authors.lastname: Lanceros-Méndez
authors.affiliation: CEB - Centro de Engenharia Biológica, Universidade do Minho, 4710-057 Braga, Portugal. Electronic address: kluskens@deb.uminho.pt.
authors.forename: L D
authors.initials: LD
authors.lastname: Kluskens
authors.affiliation: CEB - Centro de Engenharia Biológica, Universidade do Minho, 4710-057 Braga, Portugal.
authors.forename: M
authors.initials: M
authors.lastname: Mota
mesh_terms: D004926:Escherichia coli / Q000235:genetics* / Q000378:metabolism; D018014:Gene Transfer Techniques; D010957:Plasmids / Q000235:genetics* / Q000378:metabolism; D014169:Transformation, Bacterial
keywords: E. coli; Sepiolite; Transformation; Yoshida effect