Highly efficient CRISPR/Cas9-mediated targeted mutagenesis of multiple genes in Populus.
Identifieur interne : 001D12 ( Main/Exploration ); précédent : 001D11; suivant : 001D13Highly efficient CRISPR/Cas9-mediated targeted mutagenesis of multiple genes in Populus.
Auteurs : Ting-Ting Liu [République populaire de Chine] ; Di Fan [République populaire de Chine] ; Ling-Yu Ran [République populaire de Chine] ; Yuan-Zhong Jiang [République populaire de Chine] ; Rui Liu [République populaire de Chine] ; Ke-Ming Luo [République populaire de Chine]Source :
- Yi chuan = Hereditas [ 0253-9772 ] ; 2015.
Descripteurs français
- KwdFr :
- Analyse de mutations d'ADN (MeSH), Ciblage de gène (méthodes), Génie génétique (méthodes), Mutagenèse (MeSH), Mutation (MeSH), Oxidoreductases (génétique), Oxidoreductases (métabolisme), Populus (enzymologie), Populus (génétique), Protéines végétales (génétique), Protéines végétales (métabolisme), Reproductibilité des résultats (MeSH), Réaction de polymérisation en chaîne (MeSH), Similitude de séquences d'acides nucléiques (MeSH), Systèmes CRISPR-Cas (MeSH), Séquence nucléotidique (MeSH).
- MESH :
- enzymologie : Populus.
- génétique : Oxidoreductases, Populus, Protéines végétales.
- métabolisme : Oxidoreductases, Protéines végétales.
- méthodes : Ciblage de gène, Génie génétique.
- Analyse de mutations d'ADN, Mutagenèse, Mutation, Reproductibilité des résultats, Réaction de polymérisation en chaîne, Similitude de séquences d'acides nucléiques, Systèmes CRISPR-Cas, Séquence nucléotidique.
English descriptors
- KwdEn :
- Base Sequence (MeSH), CRISPR-Cas Systems (MeSH), DNA Mutational Analysis (MeSH), Gene Targeting (methods), Genetic Engineering (methods), Mutagenesis (MeSH), Mutation (MeSH), Oxidoreductases (genetics), Oxidoreductases (metabolism), Plant Proteins (genetics), Plant Proteins (metabolism), Polymerase Chain Reaction (MeSH), Populus (enzymology), Populus (genetics), Reproducibility of Results (MeSH), Sequence Homology, Nucleic Acid (MeSH).
- MESH :
- chemical , genetics : Oxidoreductases, Plant Proteins.
- chemical , metabolism : Oxidoreductases, Plant Proteins.
- enzymology : Populus.
- genetics : Populus.
- methods : Gene Targeting, Genetic Engineering.
- Base Sequence, CRISPR-Cas Systems, DNA Mutational Analysis, Mutagenesis, Mutation, Polymerase Chain Reaction, Reproducibility of Results, Sequence Homology, Nucleic Acid.
Abstract
The typeⅡCRISPR/Cas9 system (Clustered regularly interspaced short palindromic repeats /CRISPR-associated 9) has been widely used in bacteria, yeast, animals and plants as a targeted genome editing technique. In previous work, we have successfully knocked out the endogenous phytoene dehydrogenase (PDS) gene in Populus tomentosa Carr. using this system. To study the effect of target design on the efficiency of CRISPR/Cas9-mediated gene knockout in Populus, we analyzed the efficiency of mutagenesis using different single-guide RNA (sgRNA) that target PDS DNA sequence. We found that mismatches between the sgRNA and the target DNA resulted in decreased efficiency of mutagenesis and even failed mutagenesis. Moreover, complementarity between the 3' end nucleotide of sgRNA and target DNA is especially crucial for efficient mutagenesis. Further sequencing analysis showed that two PDS homologs in Populus, PtPDS1 and PtPDS2, could be knocked out simultaneously using this system with 86.4% and 50% efficiency, respectively. These results indicated the possibility of introducing mutations in two or more endogenous genes efficiently and obtaining multi-mutant strains of Populus using this system. We have indeed generated several knockout mutants of transcription factors and structural genes in Populus, which establishes a foundation for future studies of gene function and genetic improvement of Populus.
DOI: 10.16288/j.yczz.15-303
PubMed: 26496757
Affiliations:
Links toward previous steps (curation, corpus...)
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of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Liu, Rui" sort="Liu, Rui" uniqKey="Liu R" first="Rui" last="Liu">Rui Liu</name><affiliation wicri:level="1"><nlm:affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Luo, Ke Ming" sort="Luo, Ke Ming" uniqKey="Luo K" first="Ke-Ming" 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Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Ran, Ling Yu" sort="Ran, Ling Yu" uniqKey="Ran L" first="Ling-Yu" last="Ran">Ling-Yu Ran</name><affiliation wicri:level="1"><nlm:affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Jiang, Yuan Zhong" sort="Jiang, Yuan Zhong" uniqKey="Jiang Y" first="Yuan-Zhong" last="Jiang">Yuan-Zhong Jiang</name><affiliation wicri:level="1"><nlm:affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Liu, Rui" sort="Liu, Rui" uniqKey="Liu R" first="Rui" last="Liu">Rui Liu</name><affiliation wicri:level="1"><nlm:affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author><author><name sortKey="Luo, Ke Ming" sort="Luo, Ke Ming" uniqKey="Luo K" first="Ke-Ming" last="Luo">Ke-Ming Luo</name><affiliation wicri:level="1"><nlm:affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715</wicri:regionArea><wicri:noRegion>Chongqing 400715</wicri:noRegion></affiliation></author></analytic><series><title level="j">Yi chuan = Hereditas</title><idno type="ISSN">0253-9772</idno><imprint><date when="2015" type="published">2015</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Base Sequence (MeSH)</term><term>CRISPR-Cas Systems (MeSH)</term><term>DNA Mutational Analysis (MeSH)</term><term>Gene Targeting (methods)</term><term>Genetic Engineering (methods)</term><term>Mutagenesis (MeSH)</term><term>Mutation (MeSH)</term><term>Oxidoreductases (genetics)</term><term>Oxidoreductases (metabolism)</term><term>Plant Proteins (genetics)</term><term>Plant Proteins (metabolism)</term><term>Polymerase Chain Reaction (MeSH)</term><term>Populus (enzymology)</term><term>Populus (genetics)</term><term>Reproducibility of Results (MeSH)</term><term>Sequence Homology, Nucleic Acid (MeSH)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>Analyse de mutations d'ADN (MeSH)</term><term>Ciblage de gène (méthodes)</term><term>Génie génétique (méthodes)</term><term>Mutagenèse (MeSH)</term><term>Mutation (MeSH)</term><term>Oxidoreductases (génétique)</term><term>Oxidoreductases (métabolisme)</term><term>Populus (enzymologie)</term><term>Populus (génétique)</term><term>Protéines végétales (génétique)</term><term>Protéines végétales (métabolisme)</term><term>Reproductibilité des résultats (MeSH)</term><term>Réaction de polymérisation en chaîne (MeSH)</term><term>Similitude de séquences d'acides nucléiques (MeSH)</term><term>Systèmes CRISPR-Cas (MeSH)</term><term>Séquence nucléotidique (MeSH)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>Oxidoreductases</term><term>Plant Proteins</term></keywords><keywords scheme="MESH" type="chemical" qualifier="metabolism" xml:lang="en"><term>Oxidoreductases</term><term>Plant Proteins</term></keywords><keywords scheme="MESH" qualifier="enzymologie" xml:lang="fr"><term>Populus</term></keywords><keywords scheme="MESH" qualifier="enzymology" xml:lang="en"><term>Populus</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Populus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>Oxidoreductases</term><term>Populus</term><term>Protéines végétales</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Gene Targeting</term><term>Genetic Engineering</term></keywords><keywords scheme="MESH" qualifier="métabolisme" xml:lang="fr"><term>Oxidoreductases</term><term>Protéines végétales</term></keywords><keywords scheme="MESH" qualifier="méthodes" xml:lang="fr"><term>Ciblage de gène</term><term>Génie génétique</term></keywords><keywords scheme="MESH" xml:lang="en"><term>Base Sequence</term><term>CRISPR-Cas Systems</term><term>DNA Mutational Analysis</term><term>Mutagenesis</term><term>Mutation</term><term>Polymerase Chain Reaction</term><term>Reproducibility of Results</term><term>Sequence Homology, Nucleic Acid</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Analyse de mutations d'ADN</term><term>Mutagenèse</term><term>Mutation</term><term>Reproductibilité des résultats</term><term>Réaction de polymérisation en chaîne</term><term>Similitude de séquences d'acides nucléiques</term><term>Systèmes CRISPR-Cas</term><term>Séquence nucléotidique</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">The typeⅡCRISPR/Cas9 system (Clustered regularly interspaced short palindromic repeats /CRISPR-associated 9) has been widely used in bacteria, yeast, animals and plants as a targeted genome editing technique. In previous work, we have successfully knocked out the endogenous phytoene dehydrogenase (PDS) gene in Populus tomentosa Carr. using this system. To study the effect of target design on the efficiency of CRISPR/Cas9-mediated gene knockout in Populus, we analyzed the efficiency of mutagenesis using different single-guide RNA (sgRNA) that target PDS DNA sequence. We found that mismatches between the sgRNA and the target DNA resulted in decreased efficiency of mutagenesis and even failed mutagenesis. Moreover, complementarity between the 3' end nucleotide of sgRNA and target DNA is especially crucial for efficient mutagenesis. Further sequencing analysis showed that two PDS homologs in Populus, PtPDS1 and PtPDS2, could be knocked out simultaneously using this system with 86.4% and 50% efficiency, respectively. These results indicated the possibility of introducing mutations in two or more endogenous genes efficiently and obtaining multi-mutant strains of Populus using this system. We have indeed generated several knockout mutants of transcription factors and structural genes in Populus, which establishes a foundation for future studies of gene function and genetic improvement of Populus.</div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">26496757</PMID><DateCompleted><Year>2015</Year><Month>12</Month><Day>17</Day></DateCompleted><DateRevised><Year>2017</Year><Month>10</Month><Day>17</Day></DateRevised><Article PubModel="Print"><Journal><ISSN IssnType="Print">0253-9772</ISSN><JournalIssue CitedMedium="Print"><Volume>37</Volume><Issue>10</Issue><PubDate><Year>2015</Year><Month>10</Month></PubDate></JournalIssue><Title>Yi chuan = Hereditas</Title><ISOAbbreviation>Yi Chuan</ISOAbbreviation></Journal><ArticleTitle>Highly efficient CRISPR/Cas9-mediated targeted mutagenesis of multiple genes in Populus.</ArticleTitle><Pagination><MedlinePgn>1044-52</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.16288/j.yczz.15-303</ELocationID><Abstract><AbstractText>The typeⅡCRISPR/Cas9 system (Clustered regularly interspaced short palindromic repeats /CRISPR-associated 9) has been widely used in bacteria, yeast, animals and plants as a targeted genome editing technique. In previous work, we have successfully knocked out the endogenous phytoene dehydrogenase (PDS) gene in Populus tomentosa Carr. using this system. To study the effect of target design on the efficiency of CRISPR/Cas9-mediated gene knockout in Populus, we analyzed the efficiency of mutagenesis using different single-guide RNA (sgRNA) that target PDS DNA sequence. We found that mismatches between the sgRNA and the target DNA resulted in decreased efficiency of mutagenesis and even failed mutagenesis. Moreover, complementarity between the 3' end nucleotide of sgRNA and target DNA is especially crucial for efficient mutagenesis. Further sequencing analysis showed that two PDS homologs in Populus, PtPDS1 and PtPDS2, could be knocked out simultaneously using this system with 86.4% and 50% efficiency, respectively. These results indicated the possibility of introducing mutations in two or more endogenous genes efficiently and obtaining multi-mutant strains of Populus using this system. We have indeed generated several knockout mutants of transcription factors and structural genes in Populus, which establishes a foundation for future studies of gene function and genetic improvement of Populus.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Liu</LastName><ForeName>Ting-ting</ForeName><Initials>TT</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Fan</LastName><ForeName>Di</ForeName><Initials>D</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Ran</LastName><ForeName>Ling-yu</ForeName><Initials>LY</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Jiang</LastName><ForeName>Yuan-zhong</ForeName><Initials>YZ</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Liu</LastName><ForeName>Rui</ForeName><Initials>R</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Luo</LastName><ForeName>Ke-ming</ForeName><Initials>KM</Initials><AffiliationInfo><Affiliation>Institute of Resources Botany, School of Life Sciences, Southwest University, Chongqing 400715, China.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList></Article><MedlineJournalInfo><Country>China</Country><MedlineTA>Yi Chuan</MedlineTA><NlmUniqueID>9436478</NlmUniqueID><ISSNLinking>0253-9772</ISSNLinking></MedlineJournalInfo><ChemicalList><Chemical><RegistryNumber>0</RegistryNumber><NameOfSubstance UI="D010940">Plant Proteins</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.-</RegistryNumber><NameOfSubstance UI="D010088">Oxidoreductases</NameOfSubstance></Chemical><Chemical><RegistryNumber>EC 1.14.99.-</RegistryNumber><NameOfSubstance UI="C022398">phytoene dehydrogenase</NameOfSubstance></Chemical></ChemicalList><CitationSubset>IM</CitationSubset><MeshHeadingList><MeshHeading><DescriptorName UI="D001483" MajorTopicYN="N">Base Sequence</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D064113" MajorTopicYN="Y">CRISPR-Cas Systems</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D004252" MajorTopicYN="N">DNA Mutational Analysis</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D018390" MajorTopicYN="N">Gene Targeting</DescriptorName><QualifierName UI="Q000379" MajorTopicYN="Y">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D005818" MajorTopicYN="N">Genetic Engineering</DescriptorName><QualifierName UI="Q000379" MajorTopicYN="N">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D016296" MajorTopicYN="N">Mutagenesis</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D009154" MajorTopicYN="Y">Mutation</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D010088" MajorTopicYN="N">Oxidoreductases</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010940" MajorTopicYN="N">Plant Proteins</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName><QualifierName UI="Q000378" MajorTopicYN="N">metabolism</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D016133" MajorTopicYN="N">Polymerase Chain Reaction</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D032107" MajorTopicYN="N">Populus</DescriptorName><QualifierName UI="Q000201" MajorTopicYN="N">enzymology</QualifierName><QualifierName UI="Q000235" MajorTopicYN="Y">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D015203" MajorTopicYN="N">Reproducibility of Results</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D012689" MajorTopicYN="N">Sequence Homology, Nucleic Acid</DescriptorName></MeshHeading></MeshHeadingList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="entrez"><Year>2015</Year><Month>10</Month><Day>27</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2015</Year><Month>10</Month><Day>27</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2015</Year><Month>12</Month><Day>19</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">26496757</ArticleId><ArticleId IdType="doi">10.16288/j.yczz.15-303</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>République populaire de Chine</li></country></list><tree><country name="République populaire de Chine"><noRegion><name sortKey="Liu, Ting Ting" sort="Liu, Ting Ting" uniqKey="Liu T" first="Ting-Ting" last="Liu">Ting-Ting Liu</name></noRegion><name sortKey="Fan, Di" sort="Fan, Di" uniqKey="Fan D" first="Di" last="Fan">Di Fan</name><name sortKey="Jiang, Yuan Zhong" sort="Jiang, Yuan Zhong" uniqKey="Jiang Y" first="Yuan-Zhong" last="Jiang">Yuan-Zhong Jiang</name><name sortKey="Liu, Rui" sort="Liu, Rui" uniqKey="Liu R" first="Rui" last="Liu">Rui Liu</name><name sortKey="Luo, Ke Ming" sort="Luo, Ke Ming" uniqKey="Luo K" first="Ke-Ming" last="Luo">Ke-Ming Luo</name><name sortKey="Ran, Ling Yu" sort="Ran, Ling Yu" uniqKey="Ran L" first="Ling-Yu" last="Ran">Ling-Yu Ran</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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