The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease

ABSTRACT: Chronic granulomatous disease (CGD) is an inherited disorder of the innate immune system characterized by a defective oxidative burst of phagocytes and subsequent impairment of their microbicidal activity. Mutations in one of the NADPH-oxidase components affect gene expression or function...

Full description

Autores:
Agudelo Flórez, Piedad
López Quintero, Juan Alvaro
Redher, Jussara
Carneiro Sampaio, Magda
Costa Carvalho, Beatriz
Grumach, Annete
Condino Neto, Antonio
Tipo de recurso:
Article of investigation
Fecha de publicación:
2004
Institución:
Universidad de Antioquia
Repositorio:
Repositorio UdeA
Idioma:
eng
OAI Identifier:
oai:bibliotecadigital.udea.edu.co:10495/25728
Acceso en línea:
http://hdl.handle.net/10495/25728
Palabra clave:
Superóxidos
Superoxides
Fagocitos
Phagocytes
Humanos
Human
Neutrófilos
Neutrophils
Estallido Respiratorio
Respiratory Burst
Rights
openAccess
License
http://creativecommons.org/licenses/by/2.5/co/
id UDEA2_02c59cdc3c893beecacc16306d3eb859
oai_identifier_str oai:bibliotecadigital.udea.edu.co:10495/25728
network_acronym_str UDEA2
network_name_str Repositorio UdeA
repository_id_str
dc.title.spa.fl_str_mv The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
title The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
spellingShingle The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
Superóxidos
Superoxides
Fagocitos
Phagocytes
Humanos
Human
Neutrófilos
Neutrophils
Estallido Respiratorio
Respiratory Burst
title_short The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
title_full The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
title_fullStr The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
title_full_unstemmed The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
title_sort The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous disease
dc.creator.fl_str_mv Agudelo Flórez, Piedad
López Quintero, Juan Alvaro
Redher, Jussara
Carneiro Sampaio, Magda
Costa Carvalho, Beatriz
Grumach, Annete
Condino Neto, Antonio
dc.contributor.author.none.fl_str_mv Agudelo Flórez, Piedad
López Quintero, Juan Alvaro
Redher, Jussara
Carneiro Sampaio, Magda
Costa Carvalho, Beatriz
Grumach, Annete
Condino Neto, Antonio
dc.subject.decs.none.fl_str_mv Superóxidos
Superoxides
Fagocitos
Phagocytes
Humanos
Human
Neutrófilos
Neutrophils
Estallido Respiratorio
Respiratory Burst
topic Superóxidos
Superoxides
Fagocitos
Phagocytes
Humanos
Human
Neutrófilos
Neutrophils
Estallido Respiratorio
Respiratory Burst
description ABSTRACT: Chronic granulomatous disease (CGD) is an inherited disorder of the innate immune system characterized by a defective oxidative burst of phagocytes and subsequent impairment of their microbicidal activity. Mutations in one of the NADPH-oxidase components affect gene expression or function of this system, leading to the phenotype of CGD. Defects in gp91-phox lead to X-linked CGD, responsible for approximately 70% of CGD cases. Investigation of the highly heterogeneous genotype of CGD patients includes mutation analysis, Northern blot or Western blot assays according to the particular case. The aim of the present study was to use reverse transcription (RT)-PCR for the analysis of molecular defects responsible for X-linked CGD in eight Brazilian patients and to assess its potential for broader application to molecular screening in CGD. Total RNA was prepared from Epstein B virus-transformed B-lymphocytes and reverse transcribed using random hexamers. The resulting cDNA was PCR-amplified by specific and overlapping pairs of primers designed to amplify three regions of the gp91-phox gene: exons 1-5, 3-9, and 7-13. This strategy detected defective gp91-phox expression in seven patients. The RT-PCR results matched clinical history, biochemical data (nitroblue tetrazolium or superoxide release assay) and available mutation analysis in four cases. In three additional cases, RT-PCR results matched clinical history and biochemical data. In another case, RT-PCR was normal despite a clinical history compatible with CGD and defective respiratory burst. We conclude that this new application of RT-PCR analysis – a simple, economical and rapid method – was appropriate for screening molecular defects in 7 of 8 X-linked CGD patients.
publishDate 2004
dc.date.issued.none.fl_str_mv 2004
dc.date.accessioned.none.fl_str_mv 2022-02-01T16:06:37Z
dc.date.available.none.fl_str_mv 2022-02-01T16:06:37Z
dc.type.spa.fl_str_mv info:eu-repo/semantics/article
dc.type.coarversion.fl_str_mv http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.hasversion.spa.fl_str_mv info:eu-repo/semantics/publishedVersion
dc.type.coar.spa.fl_str_mv http://purl.org/coar/resource_type/c_2df8fbb1
dc.type.redcol.spa.fl_str_mv https://purl.org/redcol/resource_type/ART
dc.type.local.spa.fl_str_mv Artículo de investigación
format http://purl.org/coar/resource_type/c_2df8fbb1
status_str publishedVersion
dc.identifier.issn.none.fl_str_mv 0100-879X
dc.identifier.uri.none.fl_str_mv http://hdl.handle.net/10495/25728
dc.identifier.doi.none.fl_str_mv 10.1590/S0100-879X2004000500001
dc.identifier.eissn.none.fl_str_mv 1414-431X
identifier_str_mv 0100-879X
10.1590/S0100-879X2004000500001
1414-431X
url http://hdl.handle.net/10495/25728
dc.language.iso.spa.fl_str_mv eng
language eng
dc.relation.ispartofjournalabbrev.spa.fl_str_mv Braz. J. Med. Biol. Res.
dc.rights.spa.fl_str_mv info:eu-repo/semantics/openAccess
dc.rights.uri.*.fl_str_mv http://creativecommons.org/licenses/by/2.5/co/
dc.rights.accessrights.spa.fl_str_mv http://purl.org/coar/access_right/c_abf2
dc.rights.creativecommons.spa.fl_str_mv https://creativecommons.org/licenses/by/4.0/
eu_rights_str_mv openAccess
rights_invalid_str_mv http://creativecommons.org/licenses/by/2.5/co/
http://purl.org/coar/access_right/c_abf2
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dc.format.extent.spa.fl_str_mv 10
dc.format.mimetype.spa.fl_str_mv application/pdf
dc.publisher.spa.fl_str_mv Associação Brasileira de Divulgação Científica
dc.publisher.group.spa.fl_str_mv Inmunodeficiencias Primarias
dc.publisher.place.spa.fl_str_mv São Paulo, Brasil
institution Universidad de Antioquia
bitstream.url.fl_str_mv http://bibliotecadigital.udea.edu.co/bitstream/10495/25728/2/license_rdf
http://bibliotecadigital.udea.edu.co/bitstream/10495/25728/1/AgudeloPiedad_2004_UseReverseTranscriptionPCR.pdf
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spelling Agudelo Flórez, PiedadLópez Quintero, Juan AlvaroRedher, JussaraCarneiro Sampaio, MagdaCosta Carvalho, BeatrizGrumach, AnneteCondino Neto, Antonio2022-02-01T16:06:37Z2022-02-01T16:06:37Z20040100-879Xhttp://hdl.handle.net/10495/2572810.1590/S0100-879X20040005000011414-431XABSTRACT: Chronic granulomatous disease (CGD) is an inherited disorder of the innate immune system characterized by a defective oxidative burst of phagocytes and subsequent impairment of their microbicidal activity. Mutations in one of the NADPH-oxidase components affect gene expression or function of this system, leading to the phenotype of CGD. Defects in gp91-phox lead to X-linked CGD, responsible for approximately 70% of CGD cases. Investigation of the highly heterogeneous genotype of CGD patients includes mutation analysis, Northern blot or Western blot assays according to the particular case. The aim of the present study was to use reverse transcription (RT)-PCR for the analysis of molecular defects responsible for X-linked CGD in eight Brazilian patients and to assess its potential for broader application to molecular screening in CGD. Total RNA was prepared from Epstein B virus-transformed B-lymphocytes and reverse transcribed using random hexamers. The resulting cDNA was PCR-amplified by specific and overlapping pairs of primers designed to amplify three regions of the gp91-phox gene: exons 1-5, 3-9, and 7-13. This strategy detected defective gp91-phox expression in seven patients. The RT-PCR results matched clinical history, biochemical data (nitroblue tetrazolium or superoxide release assay) and available mutation analysis in four cases. In three additional cases, RT-PCR results matched clinical history and biochemical data. In another case, RT-PCR was normal despite a clinical history compatible with CGD and defective respiratory burst. We conclude that this new application of RT-PCR analysis – a simple, economical and rapid method – was appropriate for screening molecular defects in 7 of 8 X-linked CGD patients.COL001242610application/pdfengAssociação Brasileira de Divulgação CientíficaInmunodeficiencias PrimariasSão Paulo, Brasilinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/articlehttp://purl.org/coar/resource_type/c_2df8fbb1https://purl.org/redcol/resource_type/ARTArtículo de investigaciónhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/openAccesshttp://creativecommons.org/licenses/by/2.5/co/http://purl.org/coar/access_right/c_abf2https://creativecommons.org/licenses/by/4.0/The use of reverse transcription-PCR for the diagnosis of X-linked chronic granulomatous diseaseSuperóxidosSuperoxidesFagocitosPhagocytesHumanosHumanNeutrófilosNeutrophilsEstallido RespiratorioRespiratory BurstBraz. J. Med. Biol. Res.Brazilian Journal of Medical and Biological Research625634375CC-LICENSElicense_rdflicense_rdfapplication/rdf+xml; charset=utf-8927http://bibliotecadigital.udea.edu.co/bitstream/10495/25728/2/license_rdf1646d1f6b96dbbbc38035efc9239ac9cMD52ORIGINALAgudeloPiedad_2004_UseReverseTranscriptionPCR.pdfAgudeloPiedad_2004_UseReverseTranscriptionPCR.pdfArticulo de investigaciónapplication/pdf846443http://bibliotecadigital.udea.edu.co/bitstream/10495/25728/1/AgudeloPiedad_2004_UseReverseTranscriptionPCR.pdfecaf49ffe1120737e3aae43c427dc2d8MD51LICENSElicense.txtlicense.txttext/plain; charset=utf-81748http://bibliotecadigital.udea.edu.co/bitstream/10495/25728/3/license.txt8a4605be74aa9ea9d79846c1fba20a33MD5310495/25728oai:bibliotecadigital.udea.edu.co:10495/257282022-02-01 11:06:38.15Repositorio Institucional Universidad de Antioquiaandres.perez@udea.edu.coTk9URTogUExBQ0UgWU9VUiBPV04gTElDRU5TRSBIRVJFClRoaXMgc2FtcGxlIGxpY2Vuc2UgaXMgcHJvdmlkZWQgZm9yIGluZm9ybWF0aW9uYWwgcHVycG9zZXMgb25seS4KCk5PTi1FWENMVVNJVkUgRElTVFJJQlVUSU9OIExJQ0VOU0UKCkJ5IHNpZ25pbmcgYW5kIHN1Ym1pdHRpbmcgdGhpcyBsaWNlbnNlLCB5b3UgKHRoZSBhdXRob3Iocykgb3IgY29weXJpZ2h0Cm93bmVyKSBncmFudHMgdG8gRFNwYWNlIFVuaXZlcnNpdHkgKERTVSkgdGhlIG5vbi1leGNsdXNpdmUgcmlnaHQgdG8gcmVwcm9kdWNlLAp0cmFuc2xhdGUgKGFzIGRlZmluZWQgYmVsb3cpLCBhbmQvb3IgZGlzdHJpYnV0ZSB5b3VyIHN1Ym1pc3Npb24gKGluY2x1ZGluZwp0aGUgYWJzdHJhY3QpIHdvcmxkd2lkZSBpbiBwcmludCBhbmQgZWxlY3Ryb25pYyBmb3JtYXQgYW5kIGluIGFueSBtZWRpdW0sCmluY2x1ZGluZyBidXQgbm90IGxpbWl0ZWQgdG8gYXVkaW8gb3IgdmlkZW8uCgpZb3UgYWdyZWUgdGhhdCBEU1UgbWF5LCB3aXRob3V0IGNoYW5naW5nIHRoZSBjb250ZW50LCB0cmFuc2xhdGUgdGhlCnN1Ym1pc3Npb24gdG8gYW55IG1lZGl1bSBvciBmb3JtYXQgZm9yIHRoZSBwdXJwb3NlIG9mIHByZXNlcnZhdGlvbi4KCllvdSBhbHNvIGFncmVlIHRoYXQgRFNVIG1heSBrZWVwIG1vcmUgdGhhbiBvbmUgY29weSBvZiB0aGlzIHN1Ym1pc3Npb24gZm9yCnB1cnBvc2VzIG9mIHNlY3VyaXR5LCBiYWNrLXVwIGFuZCBwcmVzZXJ2YXRpb24uCgpZb3UgcmVwcmVzZW50IHRoYXQgdGhlIHN1Ym1pc3Npb24gaXMgeW91ciBvcmlnaW5hbCB3b3JrLCBhbmQgdGhhdCB5b3UgaGF2ZQp0aGUgcmlnaHQgdG8gZ3JhbnQgdGhlIHJpZ2h0cyBjb250YWluZWQgaW4gdGhpcyBsaWNlbnNlLiBZb3UgYWxzbyByZXByZXNlbnQKdGhhdCB5b3VyIHN1Ym1pc3Npb24gZG9lcyBub3QsIHRvIHRoZSBiZXN0IG9mIHlvdXIga25vd2xlZGdlLCBpbmZyaW5nZSB1cG9uCmFueW9uZSdzIGNvcHlyaWdodC4KCklmIHRoZSBzdWJtaXNzaW9uIGNvbnRhaW5zIG1hdGVyaWFsIGZvciB3aGljaCB5b3UgZG8gbm90IGhvbGQgY29weXJpZ2h0LAp5b3UgcmVwcmVzZW50IHRoYXQgeW91IGhhdmUgb2J0YWluZWQgdGhlIHVucmVzdHJpY3RlZCBwZXJtaXNzaW9uIG9mIHRoZQpjb3B5cmlnaHQgb3duZXIgdG8gZ3JhbnQgRFNVIHRoZSByaWdodHMgcmVxdWlyZWQgYnkgdGhpcyBsaWNlbnNlLCBhbmQgdGhhdApzdWNoIHRoaXJkLXBhcnR5IG93bmVkIG1hdGVyaWFsIGlzIGNsZWFybHkgaWRlbnRpZmllZCBhbmQgYWNrbm93bGVkZ2VkCndpdGhpbiB0aGUgdGV4dCBvciBjb250ZW50IG9mIHRoZSBzdWJtaXNzaW9uLgoKSUYgVEhFIFNVQk1JU1NJT04gSVMgQkFTRUQgVVBPTiBXT1JLIFRIQVQgSEFTIEJFRU4gU1BPTlNPUkVEIE9SIFNVUFBPUlRFRApCWSBBTiBBR0VOQ1kgT1IgT1JHQU5JWkFUSU9OIE9USEVSIFRIQU4gRFNVLCBZT1UgUkVQUkVTRU5UIFRIQVQgWU9VIEhBVkUKRlVMRklMTEVEIEFOWSBSSUdIVCBPRiBSRVZJRVcgT1IgT1RIRVIgT0JMSUdBVElPTlMgUkVRVUlSRUQgQlkgU1VDSApDT05UUkFDVCBPUiBBR1JFRU1FTlQuCgpEU1Ugd2lsbCBjbGVhcmx5IGlkZW50aWZ5IHlvdXIgbmFtZShzKSBhcyB0aGUgYXV0aG9yKHMpIG9yIG93bmVyKHMpIG9mIHRoZQpzdWJtaXNzaW9uLCBhbmQgd2lsbCBub3QgbWFrZSBhbnkgYWx0ZXJhdGlvbiwgb3RoZXIgdGhhbiBhcyBhbGxvd2VkIGJ5IHRoaXMKbGljZW5zZSwgdG8geW91ciBzdWJtaXNzaW9uLgo=