Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages
Autor(a) principal: | |
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Data de Publicação: | 2014 |
Outros Autores: | , |
Tipo de documento: | Artigo |
Idioma: | eng |
Título da fonte: | Repositório Institucional da UFRGS |
Texto Completo: | http://hdl.handle.net/10183/115296 |
Resumo: | In recent years, a significant amount of sequence data (both genomic and transcriptomic) for Echinococcus spp. has been published, thereby facilitating the analysis of genes expressed during a specific stage or involved in parasite development. To perform a suitable gene expression quantification analysis, the use of validated reference genes is strongly recommended. Thus, the aim of this work was to identify suitable reference genes to allow reliable expression normalization for genes of interest in Echinococcus granulosus sensu stricto (s.s.) (G1) and Echinococcus ortleppi upon induction of the early pre-adult development. Untreated protoscoleces (PS) and pepsin-treated protoscoleces (PSP) from E. granulosus s.s. (G1) and E. ortleppi metacestode were used. The gene expression stability of eleven candidate reference genes (bTUB, NDUFV2, RPL13, TBP, CYP-1, RPII, EF-1a, bACT-1, GAPDH, ETIF4A-III and MAPK3) was assessed using geNorm, Normfinder, and RefFinder. Our qPCR data showed a good correlation with the recently published RNA-seq data. Regarding expression stability, EF-1a and TBP were the most stable genes for both species. Interestingly, bACT-1 (the most commonly used reference gene), and GAPDH and ETIF4A-III (previously identified as housekeeping genes) did not behave stably in our assay conditions. We propose the use of EF-1a as a reference gene for studies involving gene expression analysis in both PS and PSP experimental conditions for E. granulosus s.s. and E. ortleppi. To demonstrate its applicability, EF-1a was used as a normalizer gene in the relative quantification of transcripts from genes coding for antigen B subunits. The same EF-1a reference gene may be used in studies with other Echinococcus sensu lato species. This report validates suitable reference genes for species of class Cestoda, phylum Platyhelminthes, thus providing a foundation for further validation in other epidemiologically important cestode species, such as those from the Taenia genus. |
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Espínola, Sérgio MartinFerreira, Henrique BunselmeyerZaha, Arnaldo2015-04-14T01:57:50Z20141932-6203http://hdl.handle.net/10183/115296000922639In recent years, a significant amount of sequence data (both genomic and transcriptomic) for Echinococcus spp. has been published, thereby facilitating the analysis of genes expressed during a specific stage or involved in parasite development. To perform a suitable gene expression quantification analysis, the use of validated reference genes is strongly recommended. Thus, the aim of this work was to identify suitable reference genes to allow reliable expression normalization for genes of interest in Echinococcus granulosus sensu stricto (s.s.) (G1) and Echinococcus ortleppi upon induction of the early pre-adult development. Untreated protoscoleces (PS) and pepsin-treated protoscoleces (PSP) from E. granulosus s.s. (G1) and E. ortleppi metacestode were used. The gene expression stability of eleven candidate reference genes (bTUB, NDUFV2, RPL13, TBP, CYP-1, RPII, EF-1a, bACT-1, GAPDH, ETIF4A-III and MAPK3) was assessed using geNorm, Normfinder, and RefFinder. Our qPCR data showed a good correlation with the recently published RNA-seq data. Regarding expression stability, EF-1a and TBP were the most stable genes for both species. Interestingly, bACT-1 (the most commonly used reference gene), and GAPDH and ETIF4A-III (previously identified as housekeeping genes) did not behave stably in our assay conditions. We propose the use of EF-1a as a reference gene for studies involving gene expression analysis in both PS and PSP experimental conditions for E. granulosus s.s. and E. ortleppi. To demonstrate its applicability, EF-1a was used as a normalizer gene in the relative quantification of transcripts from genes coding for antigen B subunits. The same EF-1a reference gene may be used in studies with other Echinococcus sensu lato species. This report validates suitable reference genes for species of class Cestoda, phylum Platyhelminthes, thus providing a foundation for further validation in other epidemiologically important cestode species, such as those from the Taenia genus.application/pdfengPLoS ONE. San Francisco. Vol. 9, no. 7 (July 2014), e102228, 9 p.Echinococcus sppGenesValidation of suitable reference genes for expression normalization in Echinococcus spp. larval stagesEstrangeiroinfo:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/openAccessreponame:Repositório Institucional da UFRGSinstname:Universidade Federal do Rio Grande do Sul (UFRGS)instacron:UFRGSORIGINAL000922639.pdf000922639.pdfTexto completo (inglês)application/pdf818872http://www.lume.ufrgs.br/bitstream/10183/115296/1/000922639.pdf823e53cf2082e9ab20b47d72c2b9cdebMD51TEXT000922639.pdf.txt000922639.pdf.txtExtracted Texttext/plain47084http://www.lume.ufrgs.br/bitstream/10183/115296/2/000922639.pdf.txtb860fc1ad243f26f29eba453b73eae1bMD52THUMBNAIL000922639.pdf.jpg000922639.pdf.jpgGenerated Thumbnailimage/jpeg2185http://www.lume.ufrgs.br/bitstream/10183/115296/3/000922639.pdf.jpg24e1a00cf73918694d0de10b50b4399fMD5310183/1152962023-09-24 03:40:19.673791oai:www.lume.ufrgs.br:10183/115296Repositório de PublicaçõesPUBhttps://lume.ufrgs.br/oai/requestopendoar:2023-09-24T06:40:19Repositório Institucional da UFRGS - Universidade Federal do Rio Grande do Sul (UFRGS)false |
dc.title.pt_BR.fl_str_mv |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
title |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
spellingShingle |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages Espínola, Sérgio Martin Echinococcus spp Genes |
title_short |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
title_full |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
title_fullStr |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
title_full_unstemmed |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
title_sort |
Validation of suitable reference genes for expression normalization in Echinococcus spp. larval stages |
author |
Espínola, Sérgio Martin |
author_facet |
Espínola, Sérgio Martin Ferreira, Henrique Bunselmeyer Zaha, Arnaldo |
author_role |
author |
author2 |
Ferreira, Henrique Bunselmeyer Zaha, Arnaldo |
author2_role |
author author |
dc.contributor.author.fl_str_mv |
Espínola, Sérgio Martin Ferreira, Henrique Bunselmeyer Zaha, Arnaldo |
dc.subject.por.fl_str_mv |
Echinococcus spp Genes |
topic |
Echinococcus spp Genes |
description |
In recent years, a significant amount of sequence data (both genomic and transcriptomic) for Echinococcus spp. has been published, thereby facilitating the analysis of genes expressed during a specific stage or involved in parasite development. To perform a suitable gene expression quantification analysis, the use of validated reference genes is strongly recommended. Thus, the aim of this work was to identify suitable reference genes to allow reliable expression normalization for genes of interest in Echinococcus granulosus sensu stricto (s.s.) (G1) and Echinococcus ortleppi upon induction of the early pre-adult development. Untreated protoscoleces (PS) and pepsin-treated protoscoleces (PSP) from E. granulosus s.s. (G1) and E. ortleppi metacestode were used. The gene expression stability of eleven candidate reference genes (bTUB, NDUFV2, RPL13, TBP, CYP-1, RPII, EF-1a, bACT-1, GAPDH, ETIF4A-III and MAPK3) was assessed using geNorm, Normfinder, and RefFinder. Our qPCR data showed a good correlation with the recently published RNA-seq data. Regarding expression stability, EF-1a and TBP were the most stable genes for both species. Interestingly, bACT-1 (the most commonly used reference gene), and GAPDH and ETIF4A-III (previously identified as housekeeping genes) did not behave stably in our assay conditions. We propose the use of EF-1a as a reference gene for studies involving gene expression analysis in both PS and PSP experimental conditions for E. granulosus s.s. and E. ortleppi. To demonstrate its applicability, EF-1a was used as a normalizer gene in the relative quantification of transcripts from genes coding for antigen B subunits. The same EF-1a reference gene may be used in studies with other Echinococcus sensu lato species. This report validates suitable reference genes for species of class Cestoda, phylum Platyhelminthes, thus providing a foundation for further validation in other epidemiologically important cestode species, such as those from the Taenia genus. |
publishDate |
2014 |
dc.date.issued.fl_str_mv |
2014 |
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2015-04-14T01:57:50Z |
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1932-6203 |
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000922639 |
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http://hdl.handle.net/10183/115296 |
dc.language.iso.fl_str_mv |
eng |
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dc.relation.ispartof.pt_BR.fl_str_mv |
PLoS ONE. San Francisco. Vol. 9, no. 7 (July 2014), e102228, 9 p. |
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