Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course

Detalhes bibliográficos
Autor(a) principal: Santos, Tiago
Data de Publicação: 2019
Outros Autores: Pereira, Patrícia, Queiroz, João, Cruz, Carla, Sousa, Fani
Tipo de documento: Artigo
Idioma: eng
Título da fonte: Repositório Científico de Acesso Aberto de Portugal (Repositórios Cientìficos)
Texto Completo: http://hdl.handle.net/10400.6/8186
Resumo: This laboratory experiment describes the production and purification of plasmid DNA for undergraduate biochemistry and biotechnology courses. This experiment performed in a one-week period includes the protocols for plasmid pVAX1-LacZ production in Escherichia coli DH5α cells and subsequent purification of supercoiled pVAX1-LacZ. Firstly, the students use a growth medium that favors the replication of the plasmid resulting in a higher plasmid production during exponential growth. Afterwards, alkaline lysis is done to disrupt the bacterial cells and recover pVAX1-LacZ plasmid. Lastly, they perform the purification of pVAX1-LacZ supercoiled isoform by L-histidine chromatography, followed by agarose gel electrophoresis to characterize the separation of supercoiled isoform from contaminants. The proposed experiment provides an opportunity for students to acquire these skills that are routinely used in biochemistry and biotechnology laboratories. © 2019 International Union of Biochemistry and Molecular Biology, 47(6):638-643, 2019.
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spelling Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory courseBiochemistryBiotechnologyDNA - BacterialEscherichia coliHumansLaboratoriesPlasmidsStudentsUniversitiesCurriculumThis laboratory experiment describes the production and purification of plasmid DNA for undergraduate biochemistry and biotechnology courses. This experiment performed in a one-week period includes the protocols for plasmid pVAX1-LacZ production in Escherichia coli DH5α cells and subsequent purification of supercoiled pVAX1-LacZ. Firstly, the students use a growth medium that favors the replication of the plasmid resulting in a higher plasmid production during exponential growth. Afterwards, alkaline lysis is done to disrupt the bacterial cells and recover pVAX1-LacZ plasmid. Lastly, they perform the purification of pVAX1-LacZ supercoiled isoform by L-histidine chromatography, followed by agarose gel electrophoresis to characterize the separation of supercoiled isoform from contaminants. The proposed experiment provides an opportunity for students to acquire these skills that are routinely used in biochemistry and biotechnology laboratories. © 2019 International Union of Biochemistry and Molecular Biology, 47(6):638-643, 2019.WileyuBibliorumSantos, TiagoPereira, PatríciaQueiroz, JoãoCruz, CarlaSousa, Fani2020-01-09T17:23:31Z20192019-01-01T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10400.6/8186eng10.1002/bmb.21290metadata only accessinfo: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:RCAAP2023-12-15T09:47:59Zoai:ubibliorum.ubi.pt:10400.6/8186Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-20T00:48:34.659152Repositório 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 Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
title Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
spellingShingle Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
Santos, Tiago
Biochemistry
Biotechnology
DNA - Bacterial
Escherichia coli
Humans
Laboratories
Plasmids
Students
Universities
Curriculum
title_short Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
title_full Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
title_fullStr Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
title_full_unstemmed Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
title_sort Plasmid production and purification: An integrated experiment‐based biochemistry and biotechnology laboratory course
author Santos, Tiago
author_facet Santos, Tiago
Pereira, Patrícia
Queiroz, João
Cruz, Carla
Sousa, Fani
author_role author
author2 Pereira, Patrícia
Queiroz, João
Cruz, Carla
Sousa, Fani
author2_role author
author
author
author
dc.contributor.none.fl_str_mv uBibliorum
dc.contributor.author.fl_str_mv Santos, Tiago
Pereira, Patrícia
Queiroz, João
Cruz, Carla
Sousa, Fani
dc.subject.por.fl_str_mv Biochemistry
Biotechnology
DNA - Bacterial
Escherichia coli
Humans
Laboratories
Plasmids
Students
Universities
Curriculum
topic Biochemistry
Biotechnology
DNA - Bacterial
Escherichia coli
Humans
Laboratories
Plasmids
Students
Universities
Curriculum
description This laboratory experiment describes the production and purification of plasmid DNA for undergraduate biochemistry and biotechnology courses. This experiment performed in a one-week period includes the protocols for plasmid pVAX1-LacZ production in Escherichia coli DH5α cells and subsequent purification of supercoiled pVAX1-LacZ. Firstly, the students use a growth medium that favors the replication of the plasmid resulting in a higher plasmid production during exponential growth. Afterwards, alkaline lysis is done to disrupt the bacterial cells and recover pVAX1-LacZ plasmid. Lastly, they perform the purification of pVAX1-LacZ supercoiled isoform by L-histidine chromatography, followed by agarose gel electrophoresis to characterize the separation of supercoiled isoform from contaminants. The proposed experiment provides an opportunity for students to acquire these skills that are routinely used in biochemistry and biotechnology laboratories. © 2019 International Union of Biochemistry and Molecular Biology, 47(6):638-643, 2019.
publishDate 2019
dc.date.none.fl_str_mv 2019
2019-01-01T00:00:00Z
2020-01-09T17:23:31Z
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/10400.6/8186
url http://hdl.handle.net/10400.6/8186
dc.language.iso.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv 10.1002/bmb.21290
dc.rights.driver.fl_str_mv metadata only access
info:eu-repo/semantics/openAccess
rights_invalid_str_mv metadata only access
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.publisher.none.fl_str_mv Wiley
publisher.none.fl_str_mv Wiley
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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