Two-step process for initial capture of plasmid DNA and partial removal of RNA using aqueous two-phase systems |
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| Institution: | 1. Biochemical Recovery Group, Department of Chemical Engineering, School of Engineering, University of Birmingham, Edgbaston, Birmingham B15 2TT, UK;2. Business School Newcastle University, Citywall, Citygate, St James Boulevard, Newcastle upon Tyne NE1 4JH, UK;1. Institute of Petrochemistry and Catalysis, Russian Academy of Science, 141 Prospekt Oktyabrya, 450075, Ufa, Russian Federation;2. Department of Genetics and Fundamental Medicine, Bashkir State University, 32 Zaki Validi Street, Ufa, Bashkortostan 450043, Russian Federation;3. Department of Immunology and Human Reproductive Health, Bashkir State Medical University, 3 Lenin Street, Ufa, Bashkortostan 450003, Russian Federation;4. N. N. Vorozhtsov Novosibirsk Institute of Organic Chemistry, Siberian Branch, Russian Academy of Sciences, Lavrentjev Avenue 9, Novosibirsk 630090, Russian Federation;1. Department of Gynecology, Cancer Hospital Affiliated to Zhengzhou University, Henan Cancer Hospital, Zhengzhou 450008, People''s Republic of China;2. Pathological Department, Cancer Hospital Affiliated to Zhengzhou University, Henan Cancer Hospital, Zhengzhou 450008, People''s Republic of China;3. Department of Pathology and Pathological Physiology, Basic Medical College of Zhengzhou University, Zhengzhou 450052, People''s Republic of China;1. UMR INRA 1282 Infectiologie et Santé Publique, Université de Tours, UFR des Sciences Pharmaceutiques, 31 avenue Monge, 37200 Tours, France;2. Department of Pharmacognosy, Faculty of Pharmacy, University of Gezira, Sudan;3. Inserm UMR 1069 Nutrition, Croissance et Cancer, Université de Tours, UFR des Sciences Pharmaceutiques, 31 avenue Monge, 37200 Tours, France |
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| Abstract: | In this paper, a two-step process for initial capture of plasmid DNA (pDNA) and partial removal of RNA using polyethylene glycol (PEG) and di-potassium hydrogen phosphate aqueous two-phase systems (ATPS) has been investigated. A Kühni-type ATPS extraction column was prepared with 50 ml (12% (w/w) PEG 1450, 12% (w/w) phosphate) of stationary phase and loaded with crude mobile phase (26% (w/w) PEG 1450, 4% (w/w) phosphate and 70% (w/w) lysate) at a flow rate of 6 ml min?1 at an impeller speed of 200 rpm. The experiment was terminated after 100 min, and after complete resettling of the phases, 45 ml of stationary phase was harvested. During a subsequent second extraction step contained 18% (w/w) PEG 300 and 14% (w/w) phosphate, a proportion of RNA, which was also concentrated during the column process, was removed. It was demonstrated that the recovery of pDNA in the second bottom phase was 89.4%, which was similar to the initial recovery after column extraction (92.1%). |
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