Selection and validation of reference genes for quantitative expression analysis of miRNAs and mRNAs in Poplar.
Identifieur interne : 000722 ( Main/Exploration ); précédent : 000721; suivant : 000723Selection and validation of reference genes for quantitative expression analysis of miRNAs and mRNAs in Poplar.
Auteurs : Fang Tang [République populaire de Chine] ; Liwei Chu [République populaire de Chine] ; Wenbo Shu [République populaire de Chine] ; Xuejiao He [République populaire de Chine] ; Lijuan Wang [République populaire de Chine] ; Mengzhu Lu [République populaire de Chine]Source :
- Plant methods [ 1746-4811 ] ; 2019.
Abstract
Background
Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) is a rapid and sensitive approach to identify miRNA and protein-coding gene expression in plants. However, because of the specially designated reverse transcription and shorter PCR products, very few reference genes have been identified for the quantitative analysis of miRNA expression in plants, and different internal reference genes are needed to normalize the expression of miRNAs and mRNA genes respectively. Therefore, it is particularly important to select the suitable common reference genes for normalization of quantitative PCR of miRNA and mRNA.
Results
In this study, a modified reverse transcription PCR protocol was adopted for selecting and validating universal internal reference genes of mRNAs and miRNAs. Eight commonly used reference genes, four stably expressed novel genes in
Conclusions
The purpose of this study was to identify the most appropriate reference genes for qRT-PCR of miRNAs and mRNAs in different tissues at several developmental stages in poplar.
DOI: 10.1186/s13007-019-0420-1
PubMed: 30996729
PubMed Central: PMC6451301
Affiliations:
Links toward previous steps (curation, corpus...)
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China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation><affiliation wicri:level="1"><nlm:affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Lu, Mengzhu" sort="Lu, Mengzhu" uniqKey="Lu M" first="Mengzhu" last="Lu">Mengzhu Lu</name><affiliation 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first="Fang" last="Tang">Fang Tang</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation><affiliation wicri:level="1"><nlm:affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Chu, Liwei" sort="Chu, Liwei" uniqKey="Chu L" first="Liwei" last="Chu">Liwei Chu</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation></author><author><name sortKey="Shu, Wenbo" sort="Shu, Wenbo" uniqKey="Shu W" first="Wenbo" last="Shu">Wenbo Shu</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation><affiliation wicri:level="3"><nlm:affiliation>3Key Laboratory of Horticultural Plant Biology of Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, 430070 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>3Key Laboratory of Horticultural Plant Biology of Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan</wicri:regionArea><placeName><settlement type="city">Wuhan</settlement><region type="région">Hubei</region></placeName></affiliation></author><author><name sortKey="He, Xuejiao" sort="He, Xuejiao" uniqKey="He X" first="Xuejiao" last="He">Xuejiao He</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation></author><author><name sortKey="Wang, Lijuan" sort="Wang, Lijuan" uniqKey="Wang L" first="Lijuan" last="Wang">Lijuan Wang</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation><affiliation wicri:level="1"><nlm:affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author><author><name sortKey="Lu, Mengzhu" sort="Lu, Mengzhu" uniqKey="Lu M" first="Mengzhu" last="Lu">Mengzhu Lu</name><affiliation wicri:level="3"><nlm:affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing</wicri:regionArea><placeName><settlement type="city">Pékin</settlement></placeName></affiliation><affiliation wicri:level="1"><nlm:affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing</wicri:regionArea><wicri:noRegion>Nanjing</wicri:noRegion></affiliation></author></analytic><series><title level="j">Plant methods</title><idno type="ISSN">1746-4811</idno><imprint><date when="2019" type="published">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p><b>Background</b></p><p>Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) is a rapid and sensitive approach to identify miRNA and protein-coding gene expression in plants. However, because of the specially designated reverse transcription and shorter PCR products, very few reference genes have been identified for the quantitative analysis of miRNA expression in plants, and different internal reference genes are needed to normalize the expression of miRNAs and mRNA genes respectively. Therefore, it is particularly important to select the suitable common reference genes for normalization of quantitative PCR of miRNA and mRNA.</p></div><div type="abstract" xml:lang="en"><p><b>Results</b></p><p>In this study, a modified reverse transcription PCR protocol was adopted for selecting and validating universal internal reference genes of mRNAs and miRNAs. Eight commonly used reference genes, four stably expressed novel genes in </p></div><div type="abstract" xml:lang="en"><p><b>Conclusions</b></p><p>The purpose of this study was to identify the most appropriate reference genes for qRT-PCR of miRNAs and mRNAs in different tissues at several developmental stages in poplar. </p></div></front></TEI><pubmed><MedlineCitation Status="PubMed-not-MEDLINE" Owner="NLM"><PMID Version="1">30996729</PMID><DateRevised><Year>2020</Year><Month>09</Month><Day>29</Day></DateRevised><Article PubModel="Electronic-eCollection"><Journal><ISSN IssnType="Print">1746-4811</ISSN><JournalIssue CitedMedium="Print"><Volume>15</Volume><PubDate><Year>2019</Year></PubDate></JournalIssue><Title>Plant methods</Title><ISOAbbreviation>Plant Methods</ISOAbbreviation></Journal><ArticleTitle>Selection and validation of reference genes for quantitative expression analysis of miRNAs and mRNAs in Poplar.</ArticleTitle><Pagination><MedlinePgn>35</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1186/s13007-019-0420-1</ELocationID><Abstract><AbstractText Label="Background" NlmCategory="UNASSIGNED">Quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) is a rapid and sensitive approach to identify miRNA and protein-coding gene expression in plants. However, because of the specially designated reverse transcription and shorter PCR products, very few reference genes have been identified for the quantitative analysis of miRNA expression in plants, and different internal reference genes are needed to normalize the expression of miRNAs and mRNA genes respectively. Therefore, it is particularly important to select the suitable common reference genes for normalization of quantitative PCR of miRNA and mRNA.</AbstractText><AbstractText Label="Results" NlmCategory="UNASSIGNED">In this study, a modified reverse transcription PCR protocol was adopted for selecting and validating universal internal reference genes of mRNAs and miRNAs. Eight commonly used reference genes, four stably expressed novel genes in <i>Populus tremula</i>, three small noncoding RNAs and three conserved miRNAs were selected as candidate genes, and the stability of their expression was examined across a set of 38 tissue samples from four developmental stages of poplar clone 84K (<i>Populus alba</i> × <i>Populus glandulosa</i>). The expression stability of these candidate genes was evaluated systematically by four algorithms: geNorm, NormFinder, Bestkeeper and DeltaCt. The results showed that <i>Eukaryotic initiation factor 4A</i> III (<i>EIF4A</i>) and <i>U6</i>-<i>2</i> were suitable for samples of the callus stage; <i>U6</i>-<i>1</i> and <i>U6</i>-<i>2</i> were best for the seedling stage; <i>Protein phosphatase 2A</i>-<i>2</i> (<i>PP2A</i>-<i>2</i>) and <i>U6</i>-<i>1</i> were best for the plant stage; and <i>Protein phosphatase 2A</i>-<i>2</i> (<i>PP2A</i>-<i>2</i>) and <i>Oligouridylate binding protein 1B</i> (<i>UBP</i>) were the best reference genes in the adventitious root (AR) regeneration stage.</AbstractText><AbstractText Label="Conclusions" NlmCategory="UNASSIGNED">The purpose of this study was to identify the most appropriate reference genes for qRT-PCR of miRNAs and mRNAs in different tissues at several developmental stages in poplar. <i>U6</i>-<i>1</i>, <i>EIF4A</i> and <i>PP2A</i>-<i>2</i> were the three most appropriate reference genes for qRT-PCR normalization of miRNAs and mRNAs during the plant regeneration process, and <i>PP2A</i>-<i>2</i> and <i>UBP</i> represent the best reference genes in the AR regeneration stage of poplar. This work will benefit future studies of expression and function analysis of miRNAs and their target genes in poplar.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y" EqualContrib="Y"><LastName>Tang</LastName><ForeName>Fang</ForeName><Initials>F</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo><AffiliationInfo><Affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</Affiliation><Identifier Source="GRID">grid.410625.4</Identifier></AffiliationInfo></Author><Author ValidYN="Y" EqualContrib="Y"><LastName>Chu</LastName><ForeName>Liwei</ForeName><Initials>L</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Shu</LastName><ForeName>Wenbo</ForeName><Initials>W</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo><AffiliationInfo><Affiliation>3Key Laboratory of Horticultural Plant Biology of Ministry of Education, College of Horticulture and Forestry Sciences, Huazhong Agricultural University, Wuhan, 430070 China.</Affiliation><Identifier Source="ISNI">0000 0004 1790 4137</Identifier><Identifier Source="GRID">grid.35155.37</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>He</LastName><ForeName>Xuejiao</ForeName><Initials>X</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Wang</LastName><ForeName>Lijuan</ForeName><Initials>L</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo><AffiliationInfo><Affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</Affiliation><Identifier Source="GRID">grid.410625.4</Identifier></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Lu</LastName><ForeName>Mengzhu</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>1State Key Laboratory of Tree Genetics and Breeding, Key Laboratory of Tree Breeding and Cultivation of the National Forestry and Grassland Administration, Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091 China.</Affiliation><Identifier Source="ISNI">0000 0001 2104 9346</Identifier><Identifier Source="GRID">grid.216566.0</Identifier></AffiliationInfo><AffiliationInfo><Affiliation>2Co-Innovation Center for Sustainable Forestry in Southern China, Nanjing Forestry University, Nanjing, 210037 China.</Affiliation><Identifier Source="GRID">grid.410625.4</Identifier></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2019</Year><Month>04</Month><Day>06</Day></ArticleDate></Article><MedlineJournalInfo><Country>England</Country><MedlineTA>Plant Methods</MedlineTA><NlmUniqueID>101245798</NlmUniqueID><ISSNLinking>1746-4811</ISSNLinking></MedlineJournalInfo><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Development</Keyword><Keyword MajorTopicYN="N">MicroRNAs</Keyword><Keyword MajorTopicYN="N">Normalization</Keyword><Keyword MajorTopicYN="N">Poplar</Keyword><Keyword MajorTopicYN="N">Reference genes</Keyword><Keyword MajorTopicYN="N">mRNAs</Keyword><Keyword MajorTopicYN="N">qRT-PCR</Keyword></KeywordList><CoiStatement>The authors declare that they have no competing interests.</CoiStatement></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2018</Year><Month>11</Month><Day>17</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2019</Year><Month>04</Month><Day>01</Day></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2019</Year><Month>4</Month><Day>19</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2019</Year><Month>4</Month><Day>19</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2019</Year><Month>4</Month><Day>19</Day><Hour>6</Hour><Minute>1</Minute></PubMedPubDate></History><PublicationStatus>epublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">30996729</ArticleId><ArticleId IdType="doi">10.1186/s13007-019-0420-1</ArticleId><ArticleId IdType="pii">420</ArticleId><ArticleId IdType="pmc">PMC6451301</ArticleId></ArticleIdList><ReferenceList><Reference><Citation>EMBO J. 2000 Apr 3;19(7):1638-49</Citation><ArticleIdList><ArticleId IdType="pubmed">10747031</ArticleId></ArticleIdList></Reference><Reference><Citation>Methods. 2001 Dec;25(4):402-8</Citation><ArticleIdList><ArticleId IdType="pubmed">11846609</ArticleId></ArticleIdList></Reference><Reference><Citation>Genome Biol. 2002 Jun 18;3(7):RESEARCH0034</Citation><ArticleIdList><ArticleId IdType="pubmed">12184808</ArticleId></ArticleIdList></Reference><Reference><Citation>Plant Cell. 2003 Nov;15(11):2730-41</Citation><ArticleIdList><ArticleId 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last="Chu">Liwei Chu</name><name sortKey="He, Xuejiao" sort="He, Xuejiao" uniqKey="He X" first="Xuejiao" last="He">Xuejiao He</name><name sortKey="Lu, Mengzhu" sort="Lu, Mengzhu" uniqKey="Lu M" first="Mengzhu" last="Lu">Mengzhu Lu</name><name sortKey="Lu, Mengzhu" sort="Lu, Mengzhu" uniqKey="Lu M" first="Mengzhu" last="Lu">Mengzhu Lu</name><name sortKey="Shu, Wenbo" sort="Shu, Wenbo" uniqKey="Shu W" first="Wenbo" last="Shu">Wenbo Shu</name><name sortKey="Shu, Wenbo" sort="Shu, Wenbo" uniqKey="Shu W" first="Wenbo" last="Shu">Wenbo Shu</name><name sortKey="Tang, Fang" sort="Tang, Fang" uniqKey="Tang F" first="Fang" last="Tang">Fang Tang</name><name sortKey="Wang, Lijuan" sort="Wang, Lijuan" uniqKey="Wang L" first="Lijuan" last="Wang">Lijuan Wang</name><name sortKey="Wang, Lijuan" sort="Wang, Lijuan" uniqKey="Wang L" first="Lijuan" last="Wang">Lijuan Wang</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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