title: Dye affinity cryogels for plasmid DNA purification.
abstract: The aim of this study is to prepare megaporous dye-affinity cryogel discs for the purification of plasmid DNA (pDNA) from bacterial lysate. Poly(hydroxyethyl methacrylate) [PHEMA] cryogel discs were produced by free radical polymerization initiated by N,N,N',N'-tetramethylene diamine (TEMED) and ammonium persulfate (APS) redox pair in an ice bath. Cibacron Blue F3GA was used as an affinity ligand (loading amount: 68.9μmol/g polymer). The amount of pDNA adsorbed onto the PHEMA-Cibacron Blue F3GA cryogel discs first increased and then reached a plateau value (i.e., 32.5mg/g cryogel) at 3.0mg/mL pDNA concentration. Compared with the PHEMA cryogel (0.11mg/g cryogel), the pDNA adsorption capacity of the PHEMA-Cibacron Blue F3GA cryogel (32.4mg/g polymer) was improved significantly due to the Cibacron Blue 3GA immobilization onto the polymeric matrix. pDNA adsorption amount decreased from 11.7mg/g to 1.1mg/g with the increasing of NaCl concentration. The maximum pDNA adsorption was achieved at 4°C. The overall recovery of pDNA was calculated as 90%. The PHEMA-Cibacron Blue F3GA cryogel discs could be used five times without decreasing the pDNA adsorption capacity significantly. The results show that the PHEMA-Cibacron Blue F3GA cryogel discs promise high selectivity for pDNA.
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey.
authors.forename: Duygu
authors.initials: D
authors.lastname: Çimen
authors.affiliation: Vocational School of Gerede, Department of Chemistry Technology, Bolu, Turkey.
authors.forename: Fatma
authors.initials: F
authors.lastname: Yılmaz
authors.affiliation: Department of Biology, Molecular Biology Division, Hacettepe University, Ankara, Turkey.
authors.forename: Işık
authors.initials: I
authors.lastname: Perçin
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey.
authors.forename: Deniz
authors.initials: D
authors.lastname: Türkmen
authors.affiliation: Department of Chemistry, Biochemistry Division, Hacettepe University, Ankara, Turkey. Electronic address: denizli@hacettepe.edu.tr.
authors.forename: Adil
authors.initials: A
authors.lastname: Denizli
mesh_terms: D059985:Cryogels / Q000737:chemistry*; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification*; D004926:Escherichia coli / Q000737:chemistry*; D010957:Plasmids / Q000737:chemistry / Q000302:isolation & purification*; D011102:Polyhydroxyethyl Methacrylate / Q000737:chemistry*; D014227:Triazines / Q000737:chemistry*
chemical_list: D059985:Cryogels; D004269:DNA, Bacterial; D014227:Triazines; D011102:Polyhydroxyethyl Methacrylate; C012842:Cibacron Blue F 3GA
keywords: Affinity adsorption; Cibacron Blue F3GA; Cryogels; DNA purification; PHEMA