Reference Genes in the Pathosystem Phakopsora pachyrhizi/ Soybean Suitable for Normalization in Transcript Profiling.
Identifieur interne : 000088 ( Main/Corpus ); précédent : 000087; suivant : 000089Reference Genes in the Pathosystem Phakopsora pachyrhizi/ Soybean Suitable for Normalization in Transcript Profiling.
Auteurs : Daniela Hirschburger ; Manuel Müller ; Ralf T. Voegele ; Tobias LinkSource :
- International journal of molecular sciences [ 1422-0067 ] ; 2015.
English descriptors
- KwdEn :
- Gene Expression Regulation, Fungal (MeSH), Gene Expression Regulation, Plant (MeSH), Genes, Essential (MeSH), Host-Pathogen Interactions (MeSH), Phakopsora pachyrhizi (genetics), Phakopsora pachyrhizi (physiology), Plant Diseases (genetics), Plant Diseases (microbiology), Real-Time Polymerase Chain Reaction (MeSH), Soybeans (genetics), Soybeans (microbiology), Transcriptome (MeSH).
- MESH :
- genetics : Phakopsora pachyrhizi, Plant Diseases, Soybeans.
- microbiology : Plant Diseases, Soybeans.
- physiology : Phakopsora pachyrhizi.
- Gene Expression Regulation, Fungal, Gene Expression Regulation, Plant, Genes, Essential, Host-Pathogen Interactions, Real-Time Polymerase Chain Reaction, Transcriptome.
Abstract
Phakopsora pachyrhizi is a devastating pathogen on soybean, endangering soybean production worldwide. Use of Host Induced Gene Silencing (HIGS) and the study of effector proteins could provide novel strategies for pathogen control. For both approaches quantification of transcript abundance by RT-qPCR is essential. Suitable stable reference genes for normalization are indispensable to obtain accurate RT-qPCR results. According to the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and using algorithms geNorm and NormFinder we tested candidate reference genes from P. pachyrhizi and Glycine max for their suitability in normalization of transcript levels throughout the infection process. For P. pachyrhizi we recommend a combination of CytB and PDK or GAPDH for in planta experiments. Gene expression during in vitro stages and over the whole infection process was found to be highly unstable. Here, RPS14 and UbcE2 are ranked best by geNorm and NormFinder. Alternatively CytB that has the smallest Cq range (Cq: quantification cycle) could be used. We recommend specification of gene expression relative to the germ tube stage rather than to the resting urediospore stage. For studies omitting the resting spore and the appressorium stages a combination of Elf3 and RPS9, or PKD and GAPDH should be used. For normalization of soybean genes during rust infection Ukn2 and cons7 are recommended.
DOI: 10.3390/ijms160923057
PubMed: 26404265
PubMed Central: PMC4613351
Links to Exploration step
pubmed:26404265Le document en format XML
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Manuel.Mueller@uni-hohenheim.de.</nlm:affiliation></affiliation></author><author><name sortKey="Voegele, Ralf T" sort="Voegele, Ralf T" uniqKey="Voegele R" first="Ralf T" last="Voegele">Ralf T. Voegele</name><affiliation><nlm:affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Ralf.Voegele@uni-hohenheim.de.</nlm:affiliation></affiliation></author><author><name sortKey="Link, Tobias" sort="Link, Tobias" uniqKey="Link T" first="Tobias" last="Link">Tobias Link</name><affiliation><nlm:affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. 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Daniela.Hirschburger@uni-hohenheim.de.</nlm:affiliation></affiliation></author><author><name sortKey="Muller, Manuel" sort="Muller, Manuel" uniqKey="Muller M" first="Manuel" last="Müller">Manuel Müller</name><affiliation><nlm:affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Manuel.Mueller@uni-hohenheim.de.</nlm:affiliation></affiliation></author><author><name sortKey="Voegele, Ralf T" sort="Voegele, Ralf T" uniqKey="Voegele R" first="Ralf T" last="Voegele">Ralf T. Voegele</name><affiliation><nlm:affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Ralf.Voegele@uni-hohenheim.de.</nlm:affiliation></affiliation></author><author><name sortKey="Link, Tobias" sort="Link, Tobias" uniqKey="Link T" first="Tobias" last="Link">Tobias Link</name><affiliation><nlm:affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Tobias.Link@uni-hohenheim.de.</nlm:affiliation></affiliation></author></analytic><series><title level="j">International journal of molecular sciences</title><idno type="eISSN">1422-0067</idno><imprint><date when="2015" type="published">2015</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Gene Expression Regulation, Fungal (MeSH)</term><term>Gene Expression Regulation, Plant (MeSH)</term><term>Genes, Essential (MeSH)</term><term>Host-Pathogen Interactions (MeSH)</term><term>Phakopsora pachyrhizi (genetics)</term><term>Phakopsora pachyrhizi (physiology)</term><term>Plant Diseases (genetics)</term><term>Plant Diseases (microbiology)</term><term>Real-Time Polymerase Chain Reaction (MeSH)</term><term>Soybeans (genetics)</term><term>Soybeans (microbiology)</term><term>Transcriptome (MeSH)</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Phakopsora pachyrhizi</term><term>Plant Diseases</term><term>Soybeans</term></keywords><keywords scheme="MESH" qualifier="microbiology" xml:lang="en"><term>Plant Diseases</term><term>Soybeans</term></keywords><keywords scheme="MESH" qualifier="physiology" xml:lang="en"><term>Phakopsora pachyrhizi</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Gene Expression Regulation, Fungal</term><term>Gene Expression Regulation, Plant</term><term>Genes, Essential</term><term>Host-Pathogen Interactions</term><term>Real-Time Polymerase Chain Reaction</term><term>Transcriptome</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Phakopsora pachyrhizi is a devastating pathogen on soybean, endangering soybean production worldwide. Use of Host Induced Gene Silencing (HIGS) and the study of effector proteins could provide novel strategies for pathogen control. For both approaches quantification of transcript abundance by RT-qPCR is essential. Suitable stable reference genes for normalization are indispensable to obtain accurate RT-qPCR results. According to the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and using algorithms geNorm and NormFinder we tested candidate reference genes from P. pachyrhizi and Glycine max for their suitability in normalization of transcript levels throughout the infection process. For P. pachyrhizi we recommend a combination of CytB and PDK or GAPDH for in planta experiments. Gene expression during in vitro stages and over the whole infection process was found to be highly unstable. Here, RPS14 and UbcE2 are ranked best by geNorm and NormFinder. Alternatively CytB that has the smallest Cq range (Cq: quantification cycle) could be used. We recommend specification of gene expression relative to the germ tube stage rather than to the resting urediospore stage. For studies omitting the resting spore and the appressorium stages a combination of Elf3 and RPS9, or PKD and GAPDH should be used. For normalization of soybean genes during rust infection Ukn2 and cons7 are recommended. </div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">26404265</PMID><DateCompleted><Year>2016</Year><Month>07</Month><Day>21</Day></DateCompleted><DateRevised><Year>2018</Year><Month>11</Month><Day>13</Day></DateRevised><Article PubModel="Electronic"><Journal><ISSN IssnType="Electronic">1422-0067</ISSN><JournalIssue CitedMedium="Internet"><Volume>16</Volume><Issue>9</Issue><PubDate><Year>2015</Year><Month>Sep</Month><Day>23</Day></PubDate></JournalIssue><Title>International journal of molecular sciences</Title><ISOAbbreviation>Int J Mol Sci</ISOAbbreviation></Journal><ArticleTitle>Reference Genes in the Pathosystem Phakopsora pachyrhizi/ Soybean Suitable for Normalization in Transcript Profiling.</ArticleTitle><Pagination><MedlinePgn>23057-75</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.3390/ijms160923057</ELocationID><Abstract><AbstractText>Phakopsora pachyrhizi is a devastating pathogen on soybean, endangering soybean production worldwide. Use of Host Induced Gene Silencing (HIGS) and the study of effector proteins could provide novel strategies for pathogen control. For both approaches quantification of transcript abundance by RT-qPCR is essential. Suitable stable reference genes for normalization are indispensable to obtain accurate RT-qPCR results. According to the Minimum Information for Publication of Quantitative Real-Time PCR Experiments (MIQE) guidelines and using algorithms geNorm and NormFinder we tested candidate reference genes from P. pachyrhizi and Glycine max for their suitability in normalization of transcript levels throughout the infection process. For P. pachyrhizi we recommend a combination of CytB and PDK or GAPDH for in planta experiments. Gene expression during in vitro stages and over the whole infection process was found to be highly unstable. Here, RPS14 and UbcE2 are ranked best by geNorm and NormFinder. Alternatively CytB that has the smallest Cq range (Cq: quantification cycle) could be used. We recommend specification of gene expression relative to the germ tube stage rather than to the resting urediospore stage. For studies omitting the resting spore and the appressorium stages a combination of Elf3 and RPS9, or PKD and GAPDH should be used. For normalization of soybean genes during rust infection Ukn2 and cons7 are recommended. </AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Hirschburger</LastName><ForeName>Daniela</ForeName><Initials>D</Initials><AffiliationInfo><Affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Daniela.Hirschburger@uni-hohenheim.de.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Müller</LastName><ForeName>Manuel</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Manuel.Mueller@uni-hohenheim.de.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Voegele</LastName><ForeName>Ralf T</ForeName><Initials>RT</Initials><AffiliationInfo><Affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Ralf.Voegele@uni-hohenheim.de.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Link</LastName><ForeName>Tobias</ForeName><Initials>T</Initials><AffiliationInfo><Affiliation>Department of Phytopathology, Institute of Phytomedicine, Faculty of Agricultural Sciences, University of Hohenheim, Otto-Sander-Straße 5, 70599 Stuttgart, Germany. Tobias.Link@uni-hohenheim.de.</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>2015</Year><Month>09</Month><Day>23</Day></ArticleDate></Article><MedlineJournalInfo><Country>Switzerland</Country><MedlineTA>Int J Mol Sci</MedlineTA><NlmUniqueID>101092791</NlmUniqueID><ISSNLinking>1422-0067</ISSNLinking></MedlineJournalInfo><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D015966" MajorTopicYN="N">Gene Expression Regulation, Fungal</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D018506" MajorTopicYN="N">Gene Expression Regulation, Plant</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D020043" MajorTopicYN="N">Genes, 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