Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome
Autor(a) principal: | |
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Data de Publicação: | 2010 |
Outros Autores: | , , , , , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UNESP |
Texto Completo: | http://dx.doi.org/10.1186/1756-0500-3-150 http://hdl.handle.net/11449/71733 |
Resumo: | Background. From shotgun libraries used for the genomic sequencing of the phytopathogenic bacterium Xanthomonas axonopodis pv. citri (XAC), clones that were representative of the largest possible number of coding sequences (CDSs) were selected to create a DNA microarray platform on glass slides (XACarray). The creation of the XACarray allowed for the establishment of a tool that is capable of providing data for the analysis of global genome expression in this organism. Findings. The inserts from the selected clones were amplified by PCR with the universal oligonucleotide primers M13R and M13F. The obtained products were purified and fixed in duplicate on glass slides specific for use in DNA microarrays. The number of spots on the microarray totaled 6,144 and included 768 positive controls and 624 negative controls per slide. Validation of the platform was performed through hybridization of total DNA probes from XAC labeled with different fluorophores, Cy3 and Cy5. In this validation assay, 86% of all PCR products fixed on the glass slides were confirmed to present a hybridization signal greater than twice the standard deviation of the deviation of the global median signal-to-noise ration. Conclusions. Our validation of the XACArray platform using DNA-DNA hybridization revealed that it can be used to evaluate the expression of 2,365 individual CDSs from all major functional categories, which corresponds to 52.7% of the annotated CDSs of the XAC genome. As a proof of concept, we used this platform in a previously work to verify the absence of genomic regions that could not be detected by sequencing in related strains of Xanthomonas. © 2010 Moreira et al; licensee BioMed Central Ltd. |
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Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genomeBacteria (microorganisms)XanthomonasXanthomonas axonopodisXanthomonas axonopodis pv. citriBackground. From shotgun libraries used for the genomic sequencing of the phytopathogenic bacterium Xanthomonas axonopodis pv. citri (XAC), clones that were representative of the largest possible number of coding sequences (CDSs) were selected to create a DNA microarray platform on glass slides (XACarray). The creation of the XACarray allowed for the establishment of a tool that is capable of providing data for the analysis of global genome expression in this organism. Findings. The inserts from the selected clones were amplified by PCR with the universal oligonucleotide primers M13R and M13F. The obtained products were purified and fixed in duplicate on glass slides specific for use in DNA microarrays. The number of spots on the microarray totaled 6,144 and included 768 positive controls and 624 negative controls per slide. Validation of the platform was performed through hybridization of total DNA probes from XAC labeled with different fluorophores, Cy3 and Cy5. In this validation assay, 86% of all PCR products fixed on the glass slides were confirmed to present a hybridization signal greater than twice the standard deviation of the deviation of the global median signal-to-noise ration. Conclusions. Our validation of the XACArray platform using DNA-DNA hybridization revealed that it can be used to evaluate the expression of 2,365 individual CDSs from all major functional categories, which corresponds to 52.7% of the annotated CDSs of the XAC genome. As a proof of concept, we used this platform in a previously work to verify the absence of genomic regions that could not be detected by sequencing in related strains of Xanthomonas. © 2010 Moreira et al; licensee BioMed Central Ltd.Departamento de Cincias Biolágicas (DECBI) Instituto de Cincias Exatas e Biolágicas Universidade Federal de Ouro Preto, Ouro Preto, MGNcleo de Pesquisas em Cincias Biolágicas (NUPEB) Universidade Federal de Ouro Preto, Ouro Preto, MGDepartamento de Bioquímica Instituto de Química, Universidade de so Paulo, So Paulo, SPDepartamento de Engenharia Florestal Faculdade de Cincias Agrrias Universidade Federal Dos Vales Do Jequitinhonha e Mucuri, Diamantina, MGDepartamento de Tecnologia Faculdade de Cincias Agrrias e Veterinrias de Jaboticabal UNESP - Univ. Estadual Paulista, Jaboticabal, SPAlellyx Applied Genomics, Rua James Clerk Maxwell 320, Campinas SPDepartamento de Tecnologia Faculdade de Cincias Agrrias e Veterinrias de Jaboticabal UNESP - Univ. Estadual Paulista, Jaboticabal, SPUniversidade Federal de Ouro Preto (UFOP)Universidade de São Paulo (USP)Universidade Federal dos Vales do Jequitinhonha e Mucuri (UFVJM)Universidade Estadual Paulista (Unesp)Alellyx Applied GenomicsMoreira, Leandro MDe Laia, Marcelo L [UNESP]De Souza, Robson FZaini, Paulo ADa Silva, Ana CrDa Silva, Aline MFerro, Jesus A [UNESP]2014-05-27T11:24:42Z2014-05-27T11:24:42Z2010-06-25info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articleapplication/pdfhttp://dx.doi.org/10.1186/1756-0500-3-150BMC Research Notes, v. 3.1756-0500http://hdl.handle.net/11449/7173310.1186/1756-0500-3-1502-s2.0-779537990392-s2.0-77953799039.pdfScopusreponame:Repositório Institucional da UNESPinstname:Universidade Estadual Paulista (UNESP)instacron:UNESPengBMC Research Notes0,691info:eu-repo/semantics/openAccess2024-06-07T15:32:47Zoai:repositorio.unesp.br:11449/71733Repositório InstitucionalPUBhttp://repositorio.unesp.br/oai/requestopendoar:29462024-08-05T22:45:23.829480Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP)false |
dc.title.none.fl_str_mv |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
title |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
spellingShingle |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome Moreira, Leandro M Bacteria (microorganisms) Xanthomonas Xanthomonas axonopodis Xanthomonas axonopodis pv. citri |
title_short |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
title_full |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
title_fullStr |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
title_full_unstemmed |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
title_sort |
Development and validation of a Xanthomonas axonopodis pv. citri DNA microarray platform (XACarray) generated from the shotgun libraries previously used in the sequencing of this bacterial genome |
author |
Moreira, Leandro M |
author_facet |
Moreira, Leandro M De Laia, Marcelo L [UNESP] De Souza, Robson F Zaini, Paulo A Da Silva, Ana Cr Da Silva, Aline M Ferro, Jesus A [UNESP] |
author_role |
author |
author2 |
De Laia, Marcelo L [UNESP] De Souza, Robson F Zaini, Paulo A Da Silva, Ana Cr Da Silva, Aline M Ferro, Jesus A [UNESP] |
author2_role |
author author author author author author |
dc.contributor.none.fl_str_mv |
Universidade Federal de Ouro Preto (UFOP) Universidade de São Paulo (USP) Universidade Federal dos Vales do Jequitinhonha e Mucuri (UFVJM) Universidade Estadual Paulista (Unesp) Alellyx Applied Genomics |
dc.contributor.author.fl_str_mv |
Moreira, Leandro M De Laia, Marcelo L [UNESP] De Souza, Robson F Zaini, Paulo A Da Silva, Ana Cr Da Silva, Aline M Ferro, Jesus A [UNESP] |
dc.subject.por.fl_str_mv |
Bacteria (microorganisms) Xanthomonas Xanthomonas axonopodis Xanthomonas axonopodis pv. citri |
topic |
Bacteria (microorganisms) Xanthomonas Xanthomonas axonopodis Xanthomonas axonopodis pv. citri |
description |
Background. From shotgun libraries used for the genomic sequencing of the phytopathogenic bacterium Xanthomonas axonopodis pv. citri (XAC), clones that were representative of the largest possible number of coding sequences (CDSs) were selected to create a DNA microarray platform on glass slides (XACarray). The creation of the XACarray allowed for the establishment of a tool that is capable of providing data for the analysis of global genome expression in this organism. Findings. The inserts from the selected clones were amplified by PCR with the universal oligonucleotide primers M13R and M13F. The obtained products were purified and fixed in duplicate on glass slides specific for use in DNA microarrays. The number of spots on the microarray totaled 6,144 and included 768 positive controls and 624 negative controls per slide. Validation of the platform was performed through hybridization of total DNA probes from XAC labeled with different fluorophores, Cy3 and Cy5. In this validation assay, 86% of all PCR products fixed on the glass slides were confirmed to present a hybridization signal greater than twice the standard deviation of the deviation of the global median signal-to-noise ration. Conclusions. Our validation of the XACArray platform using DNA-DNA hybridization revealed that it can be used to evaluate the expression of 2,365 individual CDSs from all major functional categories, which corresponds to 52.7% of the annotated CDSs of the XAC genome. As a proof of concept, we used this platform in a previously work to verify the absence of genomic regions that could not be detected by sequencing in related strains of Xanthomonas. © 2010 Moreira et al; licensee BioMed Central Ltd. |
publishDate |
2010 |
dc.date.none.fl_str_mv |
2010-06-25 2014-05-27T11:24:42Z 2014-05-27T11:24:42Z |
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://dx.doi.org/10.1186/1756-0500-3-150 BMC Research Notes, v. 3. 1756-0500 http://hdl.handle.net/11449/71733 10.1186/1756-0500-3-150 2-s2.0-77953799039 2-s2.0-77953799039.pdf |
url |
http://dx.doi.org/10.1186/1756-0500-3-150 http://hdl.handle.net/11449/71733 |
identifier_str_mv |
BMC Research Notes, v. 3. 1756-0500 10.1186/1756-0500-3-150 2-s2.0-77953799039 2-s2.0-77953799039.pdf |
dc.language.iso.fl_str_mv |
eng |
language |
eng |
dc.relation.none.fl_str_mv |
BMC Research Notes 0,691 |
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.source.none.fl_str_mv |
Scopus reponame:Repositório Institucional da UNESP instname:Universidade Estadual Paulista (UNESP) instacron:UNESP |
instname_str |
Universidade Estadual Paulista (UNESP) |
instacron_str |
UNESP |
institution |
UNESP |
reponame_str |
Repositório Institucional da UNESP |
collection |
Repositório Institucional da UNESP |
repository.name.fl_str_mv |
Repositório Institucional da UNESP - Universidade Estadual Paulista (UNESP) |
repository.mail.fl_str_mv |
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1808129459381862400 |