Towards the design of a scalable and commercially viable technique for plasmid purification using a methacrylate monolithic stationary phase
Data(s) |
01/08/2007
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Resumo |
A monolithic stationary phase was prepared via free radical co-polymerization of ethylene glycol dimethacrylate (EDMA) and glycidyl methacrylate (GMA) with pore diameter tailored specifically for plasmid binding, retention and elution. The polymer was functionalized. with 2-chloro-N,N-diethylethylamine hydrochloride (DEAE-Cl) for anion-exchange purification of plasmid DNA (pDNA) from clarified lysate obtained from E. coli DH5α-pUC19 culture in a ribonuclease/ protease-free environment. Characterization of the monolithic resin showed a porous material, with 68% of the pores existing in the matrix having diameters above 300 nm. The final product isolated from a single-stage 5 min anion-exchange purification was a pure and homogeneous supercoiled (SC) pDNA with no gDNA, RNA and protein contamination as confirmed by ethidium bromide agarose gel electrophoresis (EtBr-AGE), enzyme restriction analysis and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. This non-toxic technique is cGMP compatible and highly scalable for production of pDNA on a commercial level. |
Identificador | |
Publicador |
John Wiley & Sons Ltd. |
Relação |
DOI:10.1002/jctb.1733 Danquah, Michael K. & Forde, Gareth M. (2007) Towards the design of a scalable and commercially viable technique for plasmid purification using a methacrylate monolithic stationary phase. Journal of Chemical Technology and Biotechnology, 82(8), pp. 752-757. |
Direitos |
Copyright 2007 Society of Chemical Industry |
Fonte |
School of Chemistry, Physics & Mechanical Engineering; Science & Engineering Faculty |
Palavras-Chave | #Anion-exchange purification #Methacrylate monolith #Plasmid DNA #Stationary phase #Ethylene glycol dimethacrylate #Glycidyl methacrylate #Plasmid #Acrylics #Copolymerization #DNA #Electrophoresis #Ethylene glycol #Free radical polymerization #Purification #2 diethylaminoethanol #cyclic GMP #ethidium bromide #free radical #methacrylic acid #methacrylic acid derivative #polymer #protein #proteinase #resin #ribonuclease #RNA #unclassified drug #agar gel electrophoresis #anion exchange #article #biotechnological procedures #cell lysate #DNA binding #DNA purification #DNA supercoiling #DNA synthesis #elution #Escherichia coli #nonhuman #particle size #polyacrylamide gel electrophoresis #polymerization #restriction mapping |
Tipo |
Journal Article |