Plasmid DNA/RNA separation by ultrafiltration: modeling and application study
Autor(a) principal: | |
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Data de Publicação: | 2014 |
Outros Autores: | , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
Texto Completo: | http://hdl.handle.net/1822/30963 |
Resumo: | RNA is one of the main soluble contaminants that needs to be separated from plasmid DNA (pDNA) during its recovery process from fermentation broths. Although significantly different in molecular size, pDNA and RNA are difficult to separate by membrane filtration due to the fact that both types of molecules are highly deformable and therefore suffer flow-induced elongation when permeating through porous membranes. The possibility of performing this separation by ultrafiltration is investigated here with the aid of a theoretical model describing the interactions between two electrically charged, flexible macromolecules that simultaneously permeate through a porous membrane. The results of the simulations, applied to pDNA and the different types of RNA present in cell lysates under typical process conditions, show that only by a careful choice of the membrane pore size and the imposed permeate flux one can achieve the required selectivity in this operation. Ultrafiltration tests using microfiltered lysates from the production of two different plasmids, pVAX1-LacZ (6050 bp) and pCAMBIA-1303 (12,361 bp), were carried out to check the validity of the theoretical predictions; the experimental results confirm these predictions and the idea that this technique can be used in practice for pDNA purification. |
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Plasmid DNA/RNA separation by ultrafiltration: modeling and application studyPlasmid DNAUltrafiltrationModelingRNAScience & TechnologyRNA is one of the main soluble contaminants that needs to be separated from plasmid DNA (pDNA) during its recovery process from fermentation broths. Although significantly different in molecular size, pDNA and RNA are difficult to separate by membrane filtration due to the fact that both types of molecules are highly deformable and therefore suffer flow-induced elongation when permeating through porous membranes. The possibility of performing this separation by ultrafiltration is investigated here with the aid of a theoretical model describing the interactions between two electrically charged, flexible macromolecules that simultaneously permeate through a porous membrane. The results of the simulations, applied to pDNA and the different types of RNA present in cell lysates under typical process conditions, show that only by a careful choice of the membrane pore size and the imposed permeate flux one can achieve the required selectivity in this operation. Ultrafiltration tests using microfiltered lysates from the production of two different plasmids, pVAX1-LacZ (6050 bp) and pCAMBIA-1303 (12,361 bp), were carried out to check the validity of the theoretical predictions; the experimental results confirm these predictions and the idea that this technique can be used in practice for pDNA purification.Fundação para a Ciência e a Tecnologia (FCT)ElsevierUniversidade do MinhoNunes, José C.Amorim, M. T. Pessoa deEscobar, Isabel C.Queiroz, J. A.Morão, António M.20142014-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/1822/30963eng0376-738810.1016/j.memsci.2014.03.036info:eu-repo/semantics/openAccessreponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãoinstacron:RCAAP2023-07-21T11:59:06ZPortal AgregadorONG |
dc.title.none.fl_str_mv |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
title |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
spellingShingle |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study Nunes, José C. Plasmid DNA Ultrafiltration Modeling RNA Science & Technology |
title_short |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
title_full |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
title_fullStr |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
title_full_unstemmed |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
title_sort |
Plasmid DNA/RNA separation by ultrafiltration: modeling and application study |
author |
Nunes, José C. |
author_facet |
Nunes, José C. Amorim, M. T. Pessoa de Escobar, Isabel C. Queiroz, J. A. Morão, António M. |
author_role |
author |
author2 |
Amorim, M. T. Pessoa de Escobar, Isabel C. Queiroz, J. A. Morão, António M. |
author2_role |
author author author author |
dc.contributor.none.fl_str_mv |
Universidade do Minho |
dc.contributor.author.fl_str_mv |
Nunes, José C. Amorim, M. T. Pessoa de Escobar, Isabel C. Queiroz, J. A. Morão, António M. |
dc.subject.por.fl_str_mv |
Plasmid DNA Ultrafiltration Modeling RNA Science & Technology |
topic |
Plasmid DNA Ultrafiltration Modeling RNA Science & Technology |
description |
RNA is one of the main soluble contaminants that needs to be separated from plasmid DNA (pDNA) during its recovery process from fermentation broths. Although significantly different in molecular size, pDNA and RNA are difficult to separate by membrane filtration due to the fact that both types of molecules are highly deformable and therefore suffer flow-induced elongation when permeating through porous membranes. The possibility of performing this separation by ultrafiltration is investigated here with the aid of a theoretical model describing the interactions between two electrically charged, flexible macromolecules that simultaneously permeate through a porous membrane. The results of the simulations, applied to pDNA and the different types of RNA present in cell lysates under typical process conditions, show that only by a careful choice of the membrane pore size and the imposed permeate flux one can achieve the required selectivity in this operation. Ultrafiltration tests using microfiltered lysates from the production of two different plasmids, pVAX1-LacZ (6050 bp) and pCAMBIA-1303 (12,361 bp), were carried out to check the validity of the theoretical predictions; the experimental results confirm these predictions and the idea that this technique can be used in practice for pDNA purification. |
publishDate |
2014 |
dc.date.none.fl_str_mv |
2014 2014-01-01T00:00:00Z |
dc.type.status.fl_str_mv |
info:eu-repo/semantics/publishedVersion |
dc.type.driver.fl_str_mv |
info:eu-repo/semantics/article |
format |
article |
status_str |
publishedVersion |
dc.identifier.uri.fl_str_mv |
http://hdl.handle.net/1822/30963 |
url |
http://hdl.handle.net/1822/30963 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
0376-7388 10.1016/j.memsci.2014.03.036 |
dc.rights.driver.fl_str_mv |
info:eu-repo/semantics/openAccess |
eu_rights_str_mv |
openAccess |
dc.format.none.fl_str_mv |
application/pdf |
dc.publisher.none.fl_str_mv |
Elsevier |
publisher.none.fl_str_mv |
Elsevier |
dc.source.none.fl_str_mv |
reponame:Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) instname:Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação instacron:RCAAP |
instname_str |
Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação |
instacron_str |
RCAAP |
institution |
RCAAP |
reponame_str |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
collection |
Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) |
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1777303656827191296 |