A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii

Autores
Frecha, Sofía Denise; Gimenez, Maria Ines; Paggi, Roberto Alejandro; de Castro, Rosana Esther; Cerletti, Micaela
Año de publicación
2025
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
The identification and characterization of promoters and regulatory elements are commonly assessed using reporter genes. However, very few of these are available for haloarchaea. In this study, we describe the construction and validation of a reporter system for the haloarchaeon Haloferax volcanii based on a modified and stable version of the carotenoid biosynthesis enzyme phytoene synthase (PSY). This tool enables the analysis of a target gene expression through direct visualization of cell pigmentation and/or the extraction and quantification of carotenoid content. The modified crtB gene encoding PSY was cloned into the pTA963 vector under two regulatable promoters previously characterized in H. volcanii: PtnaA and Pxyl, inducible with tryptophan and xylose, respectively. The construct was introduced into and expressed in a non-pigmented H. volcanii strain (ΔcrtB) under varying inducer concentrations. For both promoters, a clear increase in pigmentation was visually observed in cultures and cell pellets with increasing inducer levels. These observations were corroborated by carotenoid extraction and quantification, as well as by Western blot analysis of PSY protein levels. This work presents a robust, easy-to-use, and versatile reporter system for investigating gene expression in H. volcanii, expanding the toolkit for genetic studies in haloarchaea.
Fil: Frecha, Sofía Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Cerletti, Micaela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Materia
HALOFERAX VOLCANII
HALOARCHAEA
REPORTER SYSTEM
CAROTENOIDS
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/279731
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/279731
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcaniiFrecha, Sofía DeniseGimenez, Maria InesPaggi, Roberto Alejandrode Castro, Rosana EstherCerletti, MicaelaHALOFERAX VOLCANIIHALOARCHAEAREPORTER SYSTEMCAROTENOIDShttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1The identification and characterization of promoters and regulatory elements are commonly assessed using reporter genes. However, very few of these are available for haloarchaea. In this study, we describe the construction and validation of a reporter system for the haloarchaeon Haloferax volcanii based on a modified and stable version of the carotenoid biosynthesis enzyme phytoene synthase (PSY). This tool enables the analysis of a target gene expression through direct visualization of cell pigmentation and/or the extraction and quantification of carotenoid content. The modified crtB gene encoding PSY was cloned into the pTA963 vector under two regulatable promoters previously characterized in H. volcanii: PtnaA and Pxyl, inducible with tryptophan and xylose, respectively. The construct was introduced into and expressed in a non-pigmented H. volcanii strain (ΔcrtB) under varying inducer concentrations. For both promoters, a clear increase in pigmentation was visually observed in cultures and cell pellets with increasing inducer levels. These observations were corroborated by carotenoid extraction and quantification, as well as by Western blot analysis of PSY protein levels. This work presents a robust, easy-to-use, and versatile reporter system for investigating gene expression in H. volcanii, expanding the toolkit for genetic studies in haloarchaea.Fil: Frecha, Sofía Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaFil: Cerletti, Micaela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; ArgentinaSpringer Tokyo2025-06info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/279731Frecha, Sofía Denise; Gimenez, Maria Ines; Paggi, Roberto Alejandro; de Castro, Rosana Esther; Cerletti, Micaela; A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii; Springer Tokyo; Extremophiles; 29; 2; 6-2025; 1-101431-0651CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://link.springer.com/10.1007/s00792-025-01391-5info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2026-02-06T13:02:28Zoai:ri.conicet.gov.ar:11336/279731instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982026-02-06 13:02:28.688CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
title A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
spellingShingle A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
Frecha, Sofía Denise
HALOFERAX VOLCANII
HALOARCHAEA
REPORTER SYSTEM
CAROTENOIDS
title_short A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
title_full A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
title_fullStr A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
title_full_unstemmed A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
title_sort A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii
dc.creator.none.fl_str_mv Frecha, Sofía Denise
Gimenez, Maria Ines
Paggi, Roberto Alejandro
de Castro, Rosana Esther
Cerletti, Micaela
author Frecha, Sofía Denise
author_facet Frecha, Sofía Denise
Gimenez, Maria Ines
Paggi, Roberto Alejandro
de Castro, Rosana Esther
Cerletti, Micaela
author_role author
author2 Gimenez, Maria Ines
Paggi, Roberto Alejandro
de Castro, Rosana Esther
Cerletti, Micaela
author2_role author
author
author
author
dc.subject.none.fl_str_mv HALOFERAX VOLCANII
HALOARCHAEA
REPORTER SYSTEM
CAROTENOIDS
topic HALOFERAX VOLCANII
HALOARCHAEA
REPORTER SYSTEM
CAROTENOIDS
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv The identification and characterization of promoters and regulatory elements are commonly assessed using reporter genes. However, very few of these are available for haloarchaea. In this study, we describe the construction and validation of a reporter system for the haloarchaeon Haloferax volcanii based on a modified and stable version of the carotenoid biosynthesis enzyme phytoene synthase (PSY). This tool enables the analysis of a target gene expression through direct visualization of cell pigmentation and/or the extraction and quantification of carotenoid content. The modified crtB gene encoding PSY was cloned into the pTA963 vector under two regulatable promoters previously characterized in H. volcanii: PtnaA and Pxyl, inducible with tryptophan and xylose, respectively. The construct was introduced into and expressed in a non-pigmented H. volcanii strain (ΔcrtB) under varying inducer concentrations. For both promoters, a clear increase in pigmentation was visually observed in cultures and cell pellets with increasing inducer levels. These observations were corroborated by carotenoid extraction and quantification, as well as by Western blot analysis of PSY protein levels. This work presents a robust, easy-to-use, and versatile reporter system for investigating gene expression in H. volcanii, expanding the toolkit for genetic studies in haloarchaea.
Fil: Frecha, Sofía Denise. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Gimenez, Maria Ines. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Paggi, Roberto Alejandro. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: de Castro, Rosana Esther. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
Fil: Cerletti, Micaela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Mar del Plata. Instituto de Investigaciones Biológicas. Universidad Nacional de Mar del Plata. Facultad de Ciencias Exactas y Naturales. Instituto de Investigaciones Biológicas; Argentina
description The identification and characterization of promoters and regulatory elements are commonly assessed using reporter genes. However, very few of these are available for haloarchaea. In this study, we describe the construction and validation of a reporter system for the haloarchaeon Haloferax volcanii based on a modified and stable version of the carotenoid biosynthesis enzyme phytoene synthase (PSY). This tool enables the analysis of a target gene expression through direct visualization of cell pigmentation and/or the extraction and quantification of carotenoid content. The modified crtB gene encoding PSY was cloned into the pTA963 vector under two regulatable promoters previously characterized in H. volcanii: PtnaA and Pxyl, inducible with tryptophan and xylose, respectively. The construct was introduced into and expressed in a non-pigmented H. volcanii strain (ΔcrtB) under varying inducer concentrations. For both promoters, a clear increase in pigmentation was visually observed in cultures and cell pellets with increasing inducer levels. These observations were corroborated by carotenoid extraction and quantification, as well as by Western blot analysis of PSY protein levels. This work presents a robust, easy-to-use, and versatile reporter system for investigating gene expression in H. volcanii, expanding the toolkit for genetic studies in haloarchaea.
publishDate 2025
dc.date.none.fl_str_mv 2025-06
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/279731
Frecha, Sofía Denise; Gimenez, Maria Ines; Paggi, Roberto Alejandro; de Castro, Rosana Esther; Cerletti, Micaela; A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii; Springer Tokyo; Extremophiles; 29; 2; 6-2025; 1-10
1431-0651
CONICET Digital
CONICET
url http://hdl.handle.net/11336/279731
identifier_str_mv Frecha, Sofía Denise; Gimenez, Maria Ines; Paggi, Roberto Alejandro; de Castro, Rosana Esther; Cerletti, Micaela; A colorful new reporter system to evaluate gene expression in the archaeon Haloferax volcanii; Springer Tokyo; Extremophiles; 29; 2; 6-2025; 1-10
1431-0651
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://link.springer.com/10.1007/s00792-025-01391-5
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
application/pdf
application/pdf
application/pdf
application/pdf
dc.publisher.none.fl_str_mv Springer Tokyo
publisher.none.fl_str_mv Springer Tokyo
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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