Reference gene selection for quantitative real-time PCR in Solanum lycopersicum L. inoculated with the mycorrhizal fungus Rhizophagus irregularis.
Identifieur interne : 001025 ( Main/Exploration ); précédent : 001024; suivant : 001026Reference gene selection for quantitative real-time PCR in Solanum lycopersicum L. inoculated with the mycorrhizal fungus Rhizophagus irregularis.
Auteurs : Alejandra Fuentes [Chili] ; Javier Ortiz [Chili] ; Nicolás Saavedra [Chili] ; Luis A. Salazar [Chili] ; Claudio Meneses [Chili] ; Cesar Arriagada [Chili]Source :
- Plant physiology and biochemistry : PPB [ 1873-2690 ] ; 2016.
Descripteurs français
- KwdFr :
- MESH :
- biosynthèse : Protéines végétales.
- métabolisme : Champignons non classés, Lycopersicon esculentum, Mycorhizes.
- physiologie : Régulation de l'expression des gènes végétaux, Symbiose.
- Réaction de polymérisation en chaine en temps réel.
English descriptors
- KwdEn :
- MESH :
- chemical , biosynthesis : Plant Proteins.
- metabolism : Fungi, Unclassified, Lycopersicon esculentum, Mycorrhizae.
- physiology : Gene Expression Regulation, Plant, Symbiosis.
- Real-Time Polymerase Chain Reaction.
Abstract
The gene expression stability of candidate reference genes in the roots and leaves of Solanum lycopersicum inoculated with arbuscular mycorrhizal fungi was investigated. Eight candidate reference genes including elongation factor 1 α (EF1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), protein phosphatase 2A (PP2Acs), ribosomal protein L2 (RPL2), β-tubulin (TUB), ubiquitin (UBI) and actin (ACT) were selected, and their expression stability was assessed to determine the most stable internal reference for quantitative PCR normalization in S. lycopersicum inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. The stability of each gene was analysed in leaves and roots together and separated using the geNorm and NormFinder algorithms. Differences were detected between leaves and roots, varying among the best-ranked genes depending on the algorithm used and the tissue analysed. PGK, TUB and EF1 genes showed higher stability in roots, while EF1 and UBI had higher stability in leaves. Statistical algorithms indicated that the GAPDH gene was the least stable under the experimental conditions assayed. Then, we analysed the expression levels of the LePT4 gene, a phosphate transporter whose expression is induced by fungal colonization in host plant roots. No differences were observed when the most stable genes were used as reference genes. However, when GAPDH was used as the reference gene, we observed an overestimation of LePT4 expression. In summary, our results revealed that candidate reference genes present variable stability in S. lycopersicum arbuscular mycorrhizal symbiosis depending on the algorithm and tissue analysed. Thus, reference gene selection is an important issue for obtaining reliable results in gene expression quantification.
DOI: 10.1016/j.plaphy.2016.01.022
PubMed: 26874621
Affiliations:
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Salazar</name><affiliation wicri:level="1"><nlm:affiliation>Center of Molecular Biology and Pharmacogenetics, Scientific and Technological Bioresource Nucleus (BIOREN), Universidad de La Frontera, Temuco 4811230, Chile.</nlm:affiliation><country xml:lang="fr">Chili</country><wicri:regionArea>Center of Molecular Biology and Pharmacogenetics, Scientific and Technological Bioresource Nucleus (BIOREN), Universidad de La Frontera, Temuco 4811230</wicri:regionArea><wicri:noRegion>Temuco 4811230</wicri:noRegion></affiliation></author><author><name sortKey="Meneses, Claudio" sort="Meneses, Claudio" uniqKey="Meneses C" first="Claudio" last="Meneses">Claudio Meneses</name><affiliation wicri:level="1"><nlm:affiliation>Centro de Biotecnología Vegetal. Universidad Andres Bello, Republica 217, Santiago, Chile.</nlm:affiliation><country xml:lang="fr">Chili</country><wicri:regionArea>Centro de Biotecnología Vegetal. Universidad Andres Bello, Republica 217, Santiago</wicri:regionArea><wicri:noRegion>Santiago</wicri:noRegion></affiliation></author><author><name sortKey="Arriagada, Cesar" sort="Arriagada, Cesar" uniqKey="Arriagada C" first="Cesar" last="Arriagada">Cesar Arriagada</name><affiliation wicri:level="1"><nlm:affiliation>Laboratorio Biorremediacion, Departamento de Ciencias Forestales, Facultad de Ciencias Agropecuarias y Forestales, Universidad de La Frontera, Temuco 4811230, Chile. Electronic address: cesar.arriagada@ufrontera.cl.</nlm:affiliation><country xml:lang="fr">Chili</country><wicri:regionArea>Laboratorio Biorremediacion, Departamento de Ciencias Forestales, Facultad de Ciencias Agropecuarias y Forestales, Universidad de La Frontera, Temuco 4811230</wicri:regionArea><wicri:noRegion>Temuco 4811230</wicri:noRegion></affiliation></author></analytic><series><title level="j">Plant physiology and biochemistry : PPB</title><idno type="eISSN">1873-2690</idno><imprint><date when="2016" type="published">2016</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Fungi, Unclassified (metabolism)</term><term>Gene Expression Regulation, Plant (physiology)</term><term>Lycopersicon esculentum (metabolism)</term><term>Mycorrhizae (metabolism)</term><term>Plant Proteins (biosynthesis)</term><term>Real-Time Polymerase Chain Reaction (MeSH)</term><term>Symbiosis (physiology)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Champignons non classés (métabolisme)</term><term>Lycopersicon esculentum (métabolisme)</term><term>Mycorhizes (métabolisme)</term><term>Protéines végétales (biosynthèse)</term><term>Réaction de polymérisation en chaine en temps réel (MeSH)</term><term>Régulation de l'expression des gènes végétaux (physiologie)</term><term>Symbiose (physiologie)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="biosynthesis" xml:lang="en"><term>Plant Proteins</term></keywords><keywords scheme="MESH" qualifier="biosynthèse" xml:lang="fr"><term>Protéines végétales</term></keywords><keywords scheme="MESH" qualifier="metabolism" xml:lang="en"><term>Fungi, Unclassified</term><term>Lycopersicon esculentum</term><term>Mycorrhizae</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Champignons non classés</term><term>Lycopersicon esculentum</term><term>Mycorhizes</term></keywords><keywords scheme="MESH" qualifier="physiologie" xml:lang="fr"><term>Régulation de l'expression des gènes végétaux</term><term>Symbiose</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Gene Expression Regulation, Plant</term><term>Symbiosis</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Real-Time Polymerase Chain Reaction</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Réaction de polymérisation en chaine en temps réel</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The gene expression stability of candidate reference genes in the roots and leaves of Solanum lycopersicum inoculated with arbuscular mycorrhizal fungi was investigated. Eight candidate reference genes including elongation factor 1 α (EF1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), protein phosphatase 2A (PP2Acs), ribosomal protein L2 (RPL2), β-tubulin (TUB), ubiquitin (UBI) and actin (ACT) were selected, and their expression stability was assessed to determine the most stable internal reference for quantitative PCR normalization in S. lycopersicum inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. The stability of each gene was analysed in leaves and roots together and separated using the geNorm and NormFinder algorithms. Differences were detected between leaves and roots, varying among the best-ranked genes depending on the algorithm used and the tissue analysed. PGK, TUB and EF1 genes showed higher stability in roots, while EF1 and UBI had higher stability in leaves. Statistical algorithms indicated that the GAPDH gene was the least stable under the experimental conditions assayed. Then, we analysed the expression levels of the LePT4 gene, a phosphate transporter whose expression is induced by fungal colonization in host plant roots. No differences were observed when the most stable genes were used as reference genes. However, when GAPDH was used as the reference gene, we observed an overestimation of LePT4 expression. In summary, our results revealed that candidate reference genes present variable stability in S. lycopersicum arbuscular mycorrhizal symbiosis depending on the algorithm and tissue analysed. Thus, reference gene selection is an important issue for obtaining reliable results in gene expression quantification. </div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">26874621</PMID><DateCompleted><Year>2016</Year><Month>12</Month><Day>13</Day></DateCompleted><DateRevised><Year>2020</Year><Month>09</Month><Day>30</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1873-2690</ISSN><JournalIssue CitedMedium="Internet"><Volume>101</Volume><PubDate><Year>2016</Year><Month>Apr</Month></PubDate></JournalIssue><Title>Plant physiology and biochemistry : PPB</Title><ISOAbbreviation>Plant Physiol Biochem</ISOAbbreviation></Journal><ArticleTitle>Reference gene selection for quantitative real-time PCR in Solanum lycopersicum L. inoculated with the mycorrhizal fungus Rhizophagus irregularis.</ArticleTitle><Pagination><MedlinePgn>124-131</MedlinePgn></Pagination><ELocationID EIdType="pii" ValidYN="Y">S0981-9428(16)30021-3</ELocationID><ELocationID EIdType="doi" ValidYN="Y">10.1016/j.plaphy.2016.01.022</ELocationID><Abstract><AbstractText>The gene expression stability of candidate reference genes in the roots and leaves of Solanum lycopersicum inoculated with arbuscular mycorrhizal fungi was investigated. Eight candidate reference genes including elongation factor 1 α (EF1), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), protein phosphatase 2A (PP2Acs), ribosomal protein L2 (RPL2), β-tubulin (TUB), ubiquitin (UBI) and actin (ACT) were selected, and their expression stability was assessed to determine the most stable internal reference for quantitative PCR normalization in S. lycopersicum inoculated with the arbuscular mycorrhizal fungus Rhizophagus irregularis. The stability of each gene was analysed in leaves and roots together and separated using the geNorm and NormFinder algorithms. Differences were detected between leaves and roots, varying among the best-ranked genes depending on the algorithm used and the tissue analysed. PGK, TUB and EF1 genes showed higher stability in roots, while EF1 and UBI had higher stability in leaves. Statistical algorithms indicated that the GAPDH gene was the least stable under the experimental conditions assayed. Then, we analysed the expression levels of the LePT4 gene, a phosphate transporter whose expression is induced by fungal colonization in host plant roots. No differences were observed when the most stable genes were used as reference genes. However, when GAPDH was used as the reference gene, we observed an overestimation of LePT4 expression. In summary, our results revealed that candidate reference genes present variable stability in S. lycopersicum arbuscular mycorrhizal symbiosis depending on the algorithm and tissue analysed. Thus, reference gene selection is an important issue for obtaining reliable results in gene expression quantification. </AbstractText><CopyrightInformation>Copyright © 2016 Elsevier Masson SAS. All rights reserved.</CopyrightInformation></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Fuentes</LastName><ForeName>Alejandra</ForeName><Initials>A</Initials><AffiliationInfo><Affiliation>Laboratorio Biorremediacion, Departamento de Ciencias Forestales, Facultad de Ciencias Agropecuarias y Forestales, Universidad de La Frontera, Temuco 4811230, Chile; Programa de Doctorado en Ciencias Mención Biología Celular y Molecular Aplicada, Universidad de la Frontera, Temuco 4811230, Chile.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Ortiz</LastName><ForeName>Javier</ForeName><Initials>J</Initials><AffiliationInfo><Affiliation>Laboratorio Biorremediacion, Departamento de Ciencias Forestales, Facultad de Ciencias Agropecuarias y Forestales, Universidad de La Frontera, Temuco 4811230, Chile.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Saavedra</LastName><ForeName>Nicolás</ForeName><Initials>N</Initials><AffiliationInfo><Affiliation>Center of Molecular Biology and Pharmacogenetics, Scientific and Technological Bioresource Nucleus (BIOREN), Universidad de La Frontera, Temuco 4811230, Chile.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Salazar</LastName><ForeName>Luis A</ForeName><Initials>LA</Initials><AffiliationInfo><Affiliation>Center of Molecular Biology and Pharmacogenetics, Scientific and Technological Bioresource Nucleus (BIOREN), Universidad de La Frontera, Temuco 4811230, Chile.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Meneses</LastName><ForeName>Claudio</ForeName><Initials>C</Initials><AffiliationInfo><Affiliation>Centro de Biotecnología Vegetal. Universidad Andres Bello, Republica 217, Santiago, Chile.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Arriagada</LastName><ForeName>Cesar</ForeName><Initials>C</Initials><AffiliationInfo><Affiliation>Laboratorio Biorremediacion, Departamento de Ciencias Forestales, Facultad de Ciencias Agropecuarias y Forestales, Universidad de La Frontera, Temuco 4811230, Chile. Electronic address: cesar.arriagada@ufrontera.cl.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, Non-U.S. Gov't</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2016</Year><Month>01</Month><Day>29</Day></ArticleDate></Article><MedlineJournalInfo><Country>France</Country><MedlineTA>Plant Physiol Biochem</MedlineTA><NlmUniqueID>9882449</NlmUniqueID><ISSNLinking>0981-9428</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D010940">Plant Proteins</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D020508" MajorTopicYN="N">Fungi, Unclassified</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D018506" MajorTopicYN="N">Gene Expression Regulation, Plant</DescriptorName><QualifierName UI="Q000502" MajorTopicYN="Y">physiology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D018551" MajorTopicYN="N">Lycopersicon esculentum</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D038821" MajorTopicYN="N">Mycorrhizae</DescriptorName><QualifierName UI="Q000378" MajorTopicYN="Y">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010940" MajorTopicYN="N">Plant Proteins</DescriptorName><QualifierName UI="Q000096" MajorTopicYN="Y">biosynthesis</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D060888" MajorTopicYN="N">Real-Time Polymerase Chain Reaction</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D013559" MajorTopicYN="N">Symbiosis</DescriptorName><QualifierName UI="Q000502" MajorTopicYN="Y">physiology</QualifierName></MeshHeading></MeshHeadingList><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Arbuscular mycorrhizal fungi</Keyword><Keyword MajorTopicYN="N">Reference genes</Keyword><Keyword MajorTopicYN="N">qPCR</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2015</Year><Month>08</Month><Day>12</Day></PubMedPubDate><PubMedPubDate PubStatus="revised"><Year>2016</Year><Month>01</Month><Day>26</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2016</Year><Month>01</Month><Day>28</Day></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2016</Year><Month>2</Month><Day>15</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2016</Year><Month>2</Month><Day>15</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2016</Year><Month>12</Month><Day>15</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">26874621</ArticleId><ArticleId IdType="pii">S0981-9428(16)30021-3</ArticleId><ArticleId IdType="doi">10.1016/j.plaphy.2016.01.022</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>Chili</li></country></list><tree><country name="Chili"><noRegion><name sortKey="Fuentes, Alejandra" sort="Fuentes, Alejandra" uniqKey="Fuentes A" first="Alejandra" last="Fuentes">Alejandra Fuentes</name></noRegion><name sortKey="Arriagada, Cesar" sort="Arriagada, Cesar" uniqKey="Arriagada C" first="Cesar" last="Arriagada">Cesar Arriagada</name><name sortKey="Meneses, Claudio" sort="Meneses, Claudio" uniqKey="Meneses C" first="Claudio" last="Meneses">Claudio Meneses</name><name sortKey="Ortiz, Javier" sort="Ortiz, Javier" uniqKey="Ortiz J" first="Javier" last="Ortiz">Javier Ortiz</name><name sortKey="Saavedra, Nicolas" sort="Saavedra, Nicolas" uniqKey="Saavedra N" first="Nicolás" last="Saavedra">Nicolás Saavedra</name><name sortKey="Salazar, Luis A" sort="Salazar, Luis A" uniqKey="Salazar L" first="Luis A" last="Salazar">Luis A. 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