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https://hdl.handle.net/1822/71045
Título: | Magnesium aminoclays as plasmid delivery agents for non-competent Escherichia coli JM109 transformation |
Autor(es): | Mendes, Gabriel Pinto Kluskens, Leon Lanceros-Méndez, S. Mota, M. |
Palavras-chave: | Magnesium aminoclays Membrane permeation Microorganism transformation Nanobiohybrid complex Plasmid delivery |
Data: | Abr-2021 |
Editora: | Elsevier 1 |
Revista: | Applied Clay Science |
Citação: | Mendes, Gabriel Pinto; Kluskens, Leon D.; Lanceros-Méndez, S.; Mota, Manuel, Magnesium aminoclays as plasmid delivery agents for non-competent Escherichia coli JM109 transformation. Applied Clay Science, 204(106010), 2021 |
Resumo(s): | Magnesium aminoclays were synthesized and used to transform non-competent Escherichia coli JM109 using the exogenous plasmid pUC19. The structure determined for the Mg aminoclays is analogous to 2:1 trioctahedral smectites such as talc, with an approximate composition R8Si8Mg6O16(OH)4, where R = CH2CH2NH2, morphologically arranged in layered sheets. Mg aminoclays were employed as a cationic vehicle that enabled the passage of plasmids across the cell envelope and led to genetic modification of the host. A stock solution of 10 mg/mL of Mg aminoclays was prepared, mixed with E. coli JM109 and pUC19 plasmid, and spread over Petri dishes containing lysogeny broth (LB), isopropyl ?-D-1-thiogalactopyranoside (IPTG), 5-bromo-4-chloro-3-indolyl-?-D-galactopyranoside (X-gal), ampicillin and various concentrations of agar (14%). The transformation efficiency obtained was higher for 1% and 2% agar even though transformation also occurred at agar concentrations of 3% and 4%. The optical density of E. coli JM109 and spreading time were also adjusted, favoring transformation when cells were used in their exponential growth phase (OD600 = 1.0) and spread for 90 s. Transformation was confirmed by the growth of blue colonies in LB/IPTG/X-gal/agar Petri dishes containing ampicillin, by regrowth of biomass in liquid media containing ampicillin and by agarose gel electrophoresis of the linearized pUC19 plasmid that followed plasmidic DNA extraction from 4 blue colonies. The maximum transformation efficiency achieved was 7.0 × 103 CFU/?g pUC19. This transformation approach proved to be suitable for a convenient, cost-effective, room-temperature, risk-free and rapid transformation of non-competent E. coli JM109. |
Tipo: | Artigo |
URI: | https://hdl.handle.net/1822/71045 |
DOI: | 10.1016/j.clay.2021.106010 |
ISSN: | 0169-1317 |
Versão da editora: | https://www.sciencedirect.com/journal/applied-clay-science |
Arbitragem científica: | yes |
Acesso: | Acesso aberto |
Aparece nas coleções: | CEB - Publicações em Revistas/Séries Internacionais / Publications in International Journals/Series |
Ficheiros deste registo:
Ficheiro | Descrição | Tamanho | Formato | |
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document_54244_1.pdf | 2,67 MB | Adobe PDF | Ver/Abrir |