Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells

Detalhes bibliográficos
Autor(a) principal: Pereira, João Nuno dos Santos
Data de Publicação: 2011
Tipo de documento: Dissertação
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10362/6149
Resumo: Dissertação para obtenção do Grau de Mestre em Biotecnologia
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spelling Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cellsTransient gene expressionRecombinant protein productionCHO cellsConditioned mediumScale-upDissertação para obtenção do Grau de Mestre em BiotecnologiaTransient gene expression (TGE) allows for fast protein production in mammalian cells and has become a very important technology in the product development pipeline of biopharmaceuticals. Polyethylenimine (PEI) mediated, high-density transfections have allowed for transient processes exceeding ~300mg/L in CHO-DG44 cells. As such, the bottleneck of TGE is no more in the titers, but in the scale-up to volumes higher than 1L, because of the need for a medium exchange before transfection. It is known that if the transfection is done in a running culture, without a medium exchange (i.e in conditioned medium), the yields obtained are very low (~5 mg/L). In CHO-DG44 cells, this problem was explored from the point of view of transfection efficiency, gene delivery and transcription. A new insight is presented in this work: The low productivities are not due to a deficient gene delivery, but instead, to lower mRNA levels that we hypothesize to be related to a lower gene accessibility of the transfected plasmid. Further, the yields were improved from ~5mg/L to ~90mg/L (18-fold) by optimizing the conditions for transfecting in conditioned medium and utilizing sodium butyrate as a transcription enhancer. These results are expected to open paths for the successful scale-up of TGE.Faculdade de Ciências e TecnologiaWurm, FlorianRUNPereira, João Nuno dos Santos2011-10-19T09:27:33Z20112011-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/masterThesisapplication/pdfhttp://hdl.handle.net/10362/6149enginfo: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:RCAAP2024-03-11T03:37:11Zoai:run.unl.pt:10362/6149Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T03:16:44.487914Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informaçãofalse
dc.title.none.fl_str_mv Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
title Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
spellingShingle Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
Pereira, João Nuno dos Santos
Transient gene expression
Recombinant protein production
CHO cells
Conditioned medium
Scale-up
title_short Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
title_full Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
title_fullStr Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
title_full_unstemmed Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
title_sort Establishing a high titer transient gene expression process in conditioned media for CHO-DG44 cells
author Pereira, João Nuno dos Santos
author_facet Pereira, João Nuno dos Santos
author_role author
dc.contributor.none.fl_str_mv Wurm, Florian
RUN
dc.contributor.author.fl_str_mv Pereira, João Nuno dos Santos
dc.subject.por.fl_str_mv Transient gene expression
Recombinant protein production
CHO cells
Conditioned medium
Scale-up
topic Transient gene expression
Recombinant protein production
CHO cells
Conditioned medium
Scale-up
description Dissertação para obtenção do Grau de Mestre em Biotecnologia
publishDate 2011
dc.date.none.fl_str_mv 2011-10-19T09:27:33Z
2011
2011-01-01T00:00:00Z
dc.type.status.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.driver.fl_str_mv info:eu-repo/semantics/masterThesis
format masterThesis
status_str publishedVersion
dc.identifier.uri.fl_str_mv http://hdl.handle.net/10362/6149
url http://hdl.handle.net/10362/6149
dc.language.iso.fl_str_mv eng
language eng
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 Faculdade de Ciências e Tecnologia
publisher.none.fl_str_mv Faculdade de Ciências e Tecnologia
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)
repository.name.fl_str_mv Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos) - Agência para a Sociedade do Conhecimento (UMIC) - FCT - Sociedade da Informação
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