A possible approach for gel-based proteomic studies in recalcitrant woody plants
Autor(a) principal: | |
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Data de Publicação: | 2013 |
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/10400.18/1730 |
Resumo: | Woody plants are particularly difficult to investigate due to high phenolic, resin, and tannin contents and laborious sample preparation. In particular, protein isolation from woody plants for two-dimensional gel electrophoresis (2-DE) is challenging as secondary metabolites negatively interfere with protein extraction and separation. In this study, three protein extraction protocols, using TCA, phenol and ethanol as precipitation or extraction agents, were tested in order to select the more efficient for woody recalcitrant plant gel-based proteomics. Grapevine leaves, pine needles and cork oak ectomycorrhizal roots were used to represent woody plant species and tissues. The phenol protocol produced higher quality 2-DE gels, with increased number of resolved spots, better spot focusing and representation of all molecular mass and isoelectric point ranges tested. In order to test the compatibility of the phenol extracted proteomes with protein identification several spots were excised from the phenol gels and analyzed by mass spectrometry (MALDI-TOF/TOF). Regardless the incomplete genome/protein databases for the plant species under analysis, 49 proteins were identified by Peptide Mass Fingerprint (PMF). Proteomic data have been deposited to the ProteomeXchange with identifier PXD000224. Our results demonstrate the complexity of protein extraction from woody plant tissues and the suitability of the phenol protocol for obtaining high quality protein extracts for efficient 2-DE separation and downstream applications such as protein identification by mass spectrometry. |
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A possible approach for gel-based proteomic studies in recalcitrant woody plantsGenómica Funcional e EstruturalGrapevinePineOakEctomycorrhizal RootsProtein Extraction2-DEMass SpectrometryWoody plants are particularly difficult to investigate due to high phenolic, resin, and tannin contents and laborious sample preparation. In particular, protein isolation from woody plants for two-dimensional gel electrophoresis (2-DE) is challenging as secondary metabolites negatively interfere with protein extraction and separation. In this study, three protein extraction protocols, using TCA, phenol and ethanol as precipitation or extraction agents, were tested in order to select the more efficient for woody recalcitrant plant gel-based proteomics. Grapevine leaves, pine needles and cork oak ectomycorrhizal roots were used to represent woody plant species and tissues. The phenol protocol produced higher quality 2-DE gels, with increased number of resolved spots, better spot focusing and representation of all molecular mass and isoelectric point ranges tested. In order to test the compatibility of the phenol extracted proteomes with protein identification several spots were excised from the phenol gels and analyzed by mass spectrometry (MALDI-TOF/TOF). Regardless the incomplete genome/protein databases for the plant species under analysis, 49 proteins were identified by Peptide Mass Fingerprint (PMF). Proteomic data have been deposited to the ProteomeXchange with identifier PXD000224. Our results demonstrate the complexity of protein extraction from woody plant tissues and the suitability of the phenol protocol for obtaining high quality protein extracts for efficient 2-DE separation and downstream applications such as protein identification by mass spectrometry.SpringerOpenRepositório Científico do Instituto Nacional de SaúdeSebastiana, MónicaFigueiredo, AndreiaMonteiro, FilipaMartins, JoanaFranco, CatarinaCoelho, Ana VarelaVaz, FátimaSimões, TâniaPenque, DeborahPais, Maria SaloméFerreira, Silvia2013-09-25T13:16:12Z2013-05-082013-05-08T00:00:00Zinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://hdl.handle.net/10400.18/1730engSpringerplus. 2013 May 8;2(1):210. Print 2013 Dec.2193-1801doi:10.1186/2193-1801-2-210info: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:RCAAP2023-07-20T15:38:53Zoai:repositorio.insa.pt:10400.18/1730Portal AgregadorONGhttps://www.rcaap.pt/oai/openaireopendoar:71602024-03-19T18:36:48.481723Repositório 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 |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
title |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
spellingShingle |
A possible approach for gel-based proteomic studies in recalcitrant woody plants Sebastiana, Mónica Genómica Funcional e Estrutural Grapevine Pine Oak Ectomycorrhizal Roots Protein Extraction 2-DE Mass Spectrometry |
title_short |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
title_full |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
title_fullStr |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
title_full_unstemmed |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
title_sort |
A possible approach for gel-based proteomic studies in recalcitrant woody plants |
author |
Sebastiana, Mónica |
author_facet |
Sebastiana, Mónica Figueiredo, Andreia Monteiro, Filipa Martins, Joana Franco, Catarina Coelho, Ana Varela Vaz, Fátima Simões, Tânia Penque, Deborah Pais, Maria Salomé Ferreira, Silvia |
author_role |
author |
author2 |
Figueiredo, Andreia Monteiro, Filipa Martins, Joana Franco, Catarina Coelho, Ana Varela Vaz, Fátima Simões, Tânia Penque, Deborah Pais, Maria Salomé Ferreira, Silvia |
author2_role |
author author author author author author author author author author |
dc.contributor.none.fl_str_mv |
Repositório Científico do Instituto Nacional de Saúde |
dc.contributor.author.fl_str_mv |
Sebastiana, Mónica Figueiredo, Andreia Monteiro, Filipa Martins, Joana Franco, Catarina Coelho, Ana Varela Vaz, Fátima Simões, Tânia Penque, Deborah Pais, Maria Salomé Ferreira, Silvia |
dc.subject.por.fl_str_mv |
Genómica Funcional e Estrutural Grapevine Pine Oak Ectomycorrhizal Roots Protein Extraction 2-DE Mass Spectrometry |
topic |
Genómica Funcional e Estrutural Grapevine Pine Oak Ectomycorrhizal Roots Protein Extraction 2-DE Mass Spectrometry |
description |
Woody plants are particularly difficult to investigate due to high phenolic, resin, and tannin contents and laborious sample preparation. In particular, protein isolation from woody plants for two-dimensional gel electrophoresis (2-DE) is challenging as secondary metabolites negatively interfere with protein extraction and separation. In this study, three protein extraction protocols, using TCA, phenol and ethanol as precipitation or extraction agents, were tested in order to select the more efficient for woody recalcitrant plant gel-based proteomics. Grapevine leaves, pine needles and cork oak ectomycorrhizal roots were used to represent woody plant species and tissues. The phenol protocol produced higher quality 2-DE gels, with increased number of resolved spots, better spot focusing and representation of all molecular mass and isoelectric point ranges tested. In order to test the compatibility of the phenol extracted proteomes with protein identification several spots were excised from the phenol gels and analyzed by mass spectrometry (MALDI-TOF/TOF). Regardless the incomplete genome/protein databases for the plant species under analysis, 49 proteins were identified by Peptide Mass Fingerprint (PMF). Proteomic data have been deposited to the ProteomeXchange with identifier PXD000224. Our results demonstrate the complexity of protein extraction from woody plant tissues and the suitability of the phenol protocol for obtaining high quality protein extracts for efficient 2-DE separation and downstream applications such as protein identification by mass spectrometry. |
publishDate |
2013 |
dc.date.none.fl_str_mv |
2013-09-25T13:16:12Z 2013-05-08 2013-05-08T00:00:00Z |
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/10400.18/1730 |
url |
http://hdl.handle.net/10400.18/1730 |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
Springerplus. 2013 May 8;2(1):210. Print 2013 Dec. 2193-1801 doi:10.1186/2193-1801-2-210 |
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 |
SpringerOpen |
publisher.none.fl_str_mv |
SpringerOpen |
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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1799132100662657024 |