Determination of Reference microRNAs for Relative Quantification in Porcine Tissues

Relative quantification is the strategy of choice for processing RT-qPCR data in microRNAs (miRNAs) expression studies. Normalisation of relative quantification data is performed by using reference genes. In livestock species, such as pigs, the determination of reference miRNAs and the optimal numbe...

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Autores: Timoneda i Heredia, Oriol, Balcells Ortega, Ingrid, Córdoba, Sarai|||0000-0002-8786-9889, Castelló Farré, Anna|||0000-0001-8497-6251, Sánchez Bonastre, Armando|||0000-0001-9160-1124
Formato: artículo
Fecha de publicación:2012
País:España
Recursos:Universitat Autònoma de Barcelona
Repositorio:Dipòsit Digital de Documents de la UAB
Idioma:inglés
OAI Identifier:oai:ddd.uab.cat:141483
Acesso em linha:https://ddd.uab.cat/record/141483
https://dx.doi.org/urn:doi:10.1371/journal.pone.0044413
Access Level:acceso abierto
Palavra-chave:MicroRNAs
Skeletal muscles
Kidneys
Uterus
Ovaries
Gene expression
Swine
Algorithms
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spelling Determination of Reference microRNAs for Relative Quantification in Porcine TissuesTimoneda i Heredia, OriolBalcells Ortega, IngridCórdoba, Sarai|||0000-0002-8786-9889Castelló Farré, Anna|||0000-0001-8497-6251Sánchez Bonastre, Armando|||0000-0001-9160-1124MicroRNAsSkeletal musclesKidneysUterusOvariesGene expressionSwineAlgorithmsRelative quantification is the strategy of choice for processing RT-qPCR data in microRNAs (miRNAs) expression studies. Normalisation of relative quantification data is performed by using reference genes. In livestock species, such as pigs, the determination of reference miRNAs and the optimal number of them has not been widely studied. In this study, the stability of ten miRNAs (Ssc-let-7a, Ssc-miR-103, Ssc-miR-17-3p, Hsa-miR-25, Hsa-miR-93, Ssc-miR-106a, Ssc-miR-191, Ssc-miR-16, Ssc-miR-26a and Ssc-miR-17-5p) was investigated by RT-qPCR in different tissues (skeletal muscle, kidney, liver, ovary and uterus) and in different pig breeds (Iberian, Landrace, Large White, Meishan and Vietnamese) as variation factors. Stability values were calculated with geNorm and NormFinder algorithms obtaining high correlation between them (r2 = 0.99). The analyses showed that tissue is an important variability factor in miRNAs expression stability whereas breed is not a determinant factor. All ten miRNAs analysed had good stability values and, therefore, can be used as reference miRNAs. When all tissues were considered, miR-93 was the most stable miRNA. Dividing data set by tissues, let-7a was the most stable in skeletal muscle and ovary, miR-17-5p in kidney, miR-26a in liver and miR-103 in uterus. Moreover, the optimal number of reference miRNAs to be used for proper normalisation data was determined. It is suggested the use of five reference miRNAs (miR-93, miR-25, miR-106a, miR-17-5p and miR-26a) in multi-tissue experimental designs and the use of three reference miRNAs as the optimal number in single tissues studies (let-7a, miR-17-5p and miR-25 in skeletal muscle; miR-17-5p, miR-93 and miR-26a in kidney, miR-26a, miR-103 and let-7a in liver, let-7a, miR-25 and miR-106a in ovary and miR-103, let-7a and miR-93 in uterus). Overall, this study provides valuable information about the porcine reference miRNAs that can be used in order to perform a proper normalisation when relative quantification by RT-qPCR studies is undertaken. 22012-01-0120122012-01-01Articlehttp://purl.org/coar/resource_type/c_6501VoRhttp://purl.org/coar/version/c_970fb48d4fbd8a85info:eu-repo/semantics/articleapplication/pdfhttps://ddd.uab.cat/record/141483https://dx.doi.org/urn:doi:10.1371/journal.pone.0044413reponame:Dipòsit Digital de Documents de la UABinstname:Universitat Autònoma de BarcelonaInglésengMinisterio de Ciencia e Innovación https://doi.org/10.13039/501100004837 AGL2010-22358-C02-01Ministerio de Educación y Ciencia https://doi.org/10.13039/501100004837 AGL2007-66371-C02-01Ministerio de Educación y Ciencia https://doi.org/10.13039/501100004837 CSD2007-00036open accesshttp://purl.org/coar/access_right/c_abf2Aquest document està subjecte a una llicència d'ús Creative Commons. Es permet la reproducció total o parcial, la distribució, la comunicació pública de l'obra i la creació d'obres derivades, fins i tot amb finalitats comercials, sempre i quan es reconegui l'autoria de l'obra original.https://creativecommons.org/licenses/by/3.0/info:eu-repo/semantics/openAccessoai:ddd.uab.cat:1414832026-06-06T12:50:31Z
dc.title.none.fl_str_mv Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
title Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
spellingShingle Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
Timoneda i Heredia, Oriol
MicroRNAs
Skeletal muscles
Kidneys
Uterus
Ovaries
Gene expression
Swine
Algorithms
title_short Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
title_full Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
title_fullStr Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
title_full_unstemmed Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
title_sort Determination of Reference microRNAs for Relative Quantification in Porcine Tissues
dc.creator.none.fl_str_mv Timoneda i Heredia, Oriol
Balcells Ortega, Ingrid
Córdoba, Sarai|||0000-0002-8786-9889
Castelló Farré, Anna|||0000-0001-8497-6251
Sánchez Bonastre, Armando|||0000-0001-9160-1124
author Timoneda i Heredia, Oriol
author_facet Timoneda i Heredia, Oriol
Balcells Ortega, Ingrid
Córdoba, Sarai|||0000-0002-8786-9889
Castelló Farré, Anna|||0000-0001-8497-6251
Sánchez Bonastre, Armando|||0000-0001-9160-1124
author_role author
author2 Balcells Ortega, Ingrid
Córdoba, Sarai|||0000-0002-8786-9889
Castelló Farré, Anna|||0000-0001-8497-6251
Sánchez Bonastre, Armando|||0000-0001-9160-1124
author2_role author
author
author
author
dc.subject.none.fl_str_mv MicroRNAs
Skeletal muscles
Kidneys
Uterus
Ovaries
Gene expression
Swine
Algorithms
topic MicroRNAs
Skeletal muscles
Kidneys
Uterus
Ovaries
Gene expression
Swine
Algorithms
description Relative quantification is the strategy of choice for processing RT-qPCR data in microRNAs (miRNAs) expression studies. Normalisation of relative quantification data is performed by using reference genes. In livestock species, such as pigs, the determination of reference miRNAs and the optimal number of them has not been widely studied. In this study, the stability of ten miRNAs (Ssc-let-7a, Ssc-miR-103, Ssc-miR-17-3p, Hsa-miR-25, Hsa-miR-93, Ssc-miR-106a, Ssc-miR-191, Ssc-miR-16, Ssc-miR-26a and Ssc-miR-17-5p) was investigated by RT-qPCR in different tissues (skeletal muscle, kidney, liver, ovary and uterus) and in different pig breeds (Iberian, Landrace, Large White, Meishan and Vietnamese) as variation factors. Stability values were calculated with geNorm and NormFinder algorithms obtaining high correlation between them (r2 = 0.99). The analyses showed that tissue is an important variability factor in miRNAs expression stability whereas breed is not a determinant factor. All ten miRNAs analysed had good stability values and, therefore, can be used as reference miRNAs. When all tissues were considered, miR-93 was the most stable miRNA. Dividing data set by tissues, let-7a was the most stable in skeletal muscle and ovary, miR-17-5p in kidney, miR-26a in liver and miR-103 in uterus. Moreover, the optimal number of reference miRNAs to be used for proper normalisation data was determined. It is suggested the use of five reference miRNAs (miR-93, miR-25, miR-106a, miR-17-5p and miR-26a) in multi-tissue experimental designs and the use of three reference miRNAs as the optimal number in single tissues studies (let-7a, miR-17-5p and miR-25 in skeletal muscle; miR-17-5p, miR-93 and miR-26a in kidney, miR-26a, miR-103 and let-7a in liver, let-7a, miR-25 and miR-106a in ovary and miR-103, let-7a and miR-93 in uterus). Overall, this study provides valuable information about the porcine reference miRNAs that can be used in order to perform a proper normalisation when relative quantification by RT-qPCR studies is undertaken.
publishDate 2012
dc.date.none.fl_str_mv 2
2012-01-01
2012
2012-01-01
dc.type.none.fl_str_mv Article
http://purl.org/coar/resource_type/c_6501
VoR
http://purl.org/coar/version/c_970fb48d4fbd8a85
dc.type.openaire.fl_str_mv info:eu-repo/semantics/article
format article
dc.identifier.none.fl_str_mv https://ddd.uab.cat/record/141483
https://dx.doi.org/urn:doi:10.1371/journal.pone.0044413
url https://ddd.uab.cat/record/141483
https://dx.doi.org/urn:doi:10.1371/journal.pone.0044413
dc.language.none.fl_str_mv Inglés
eng
language_invalid_str_mv Inglés
language eng
dc.relation.none.fl_str_mv Ministerio de Ciencia e Innovación https://doi.org/10.13039/501100004837 AGL2010-22358-C02-01
Ministerio de Educación y Ciencia https://doi.org/10.13039/501100004837 AGL2007-66371-C02-01
Ministerio de Educación y Ciencia https://doi.org/10.13039/501100004837 CSD2007-00036
dc.rights.none.fl_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by/3.0/
dc.rights.openaire.fl_str_mv info:eu-repo/semantics/openAccess
rights_invalid_str_mv open access
http://purl.org/coar/access_right/c_abf2
https://creativecommons.org/licenses/by/3.0/
eu_rights_str_mv openAccess
dc.format.none.fl_str_mv application/pdf
dc.source.none.fl_str_mv reponame:Dipòsit Digital de Documents de la UAB
instname:Universitat Autònoma de Barcelona
instname_str Universitat Autònoma de Barcelona
reponame_str Dipòsit Digital de Documents de la UAB
collection Dipòsit Digital de Documents de la UAB
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