Yeast vectors for the integration/expression of any sequence at the TYR1 locus
We have constructed new yeast vectors for targeted integration and conditional expression of any sequence at the Saccharomyces cerevisiae TYR1 locus which becomes disrupted. We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectab...
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| Veröffentlicht in: | Yeast (Chichester, England) England), 2007-09, Vol.24 (9), p.761-766 |
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| container_issue | 9 |
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| container_title | Yeast (Chichester, England) |
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| creator | Mirisola, Mario G Colomba, Leonarda Gallo, Alessia Amodeo, Roberta De Leo, Giacomo |
| description | We have constructed new yeast vectors for targeted integration and conditional expression of any sequence at the Saccharomyces cerevisiae TYR1 locus which becomes disrupted. We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectable marker (uracil or leucine, depending on the vector used). This feature allows a double screening of transformed yeast cells, improving the identification of colonies with the desired chromosomal structure. The GAL10 gene promoter has been added to drive conditional expression of cloned sequences. Using these vectors, chromosomal structure verification of recombinant clones is no longer necessary, since the noise of non-homologous recombination, as well as spontaneous reversion of the selected phenotype, can easily be identified. The ability of the vector to conditionally control gene expression has been confirmed using the gene for the green fluorescent protein (GFP) as a reporter. GenBank Accession Nos EF202083-202086. Copyright © 2007 John Wiley & Sons, Ltd. |
| doi_str_mv | 10.1002/yea.1511 |
| format | Article |
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We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectable marker (uracil or leucine, depending on the vector used). This feature allows a double screening of transformed yeast cells, improving the identification of colonies with the desired chromosomal structure. The GAL10 gene promoter has been added to drive conditional expression of cloned sequences. Using these vectors, chromosomal structure verification of recombinant clones is no longer necessary, since the noise of non-homologous recombination, as well as spontaneous reversion of the selected phenotype, can easily be identified. The ability of the vector to conditionally control gene expression has been confirmed using the gene for the green fluorescent protein (GFP) as a reporter. GenBank Accession Nos EF202083-202086. 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We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectable marker (uracil or leucine, depending on the vector used). This feature allows a double screening of transformed yeast cells, improving the identification of colonies with the desired chromosomal structure. The GAL10 gene promoter has been added to drive conditional expression of cloned sequences. Using these vectors, chromosomal structure verification of recombinant clones is no longer necessary, since the noise of non-homologous recombination, as well as spontaneous reversion of the selected phenotype, can easily be identified. The ability of the vector to conditionally control gene expression has been confirmed using the gene for the green fluorescent protein (GFP) as a reporter. GenBank Accession Nos EF202083-202086. Copyright © 2007 John Wiley & Sons, Ltd.</description><subject>Base Sequence</subject><subject>DNA, Fungal - chemistry</subject><subject>DNA, Fungal - genetics</subject><subject>ends‐out gene targeting</subject><subject>expression vectors</subject><subject>Gene Expression Regulation, Fungal</subject><subject>Genes, Reporter - genetics</subject><subject>Genetic Vectors - chemistry</subject><subject>Genetic Vectors - genetics</subject><subject>Green Fluorescent Proteins - genetics</subject><subject>heterologous expression</subject><subject>Molecular Sequence Data</subject><subject>Mutagenesis, Insertional - methods</subject><subject>Polymerase Chain Reaction</subject><subject>Saccharomyces cerevisiae - genetics</subject><subject>Saccharomyces cerevisiae - growth & development</subject><subject>Sequence Analysis, DNA</subject><subject>TYR1 locus</subject><subject>Tyrosine - chemistry</subject><subject>Tyrosine - genetics</subject><subject>yeast integration vector</subject><issn>0749-503X</issn><issn>1097-0061</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2007</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNp1kE1Lw0AQQBdRbK2Cv0D36CXtzG6TTY6l1A8oCtqCPS2bZLZG2qbupmr-vakpePI0A_N4DI-xS4Q-AohBTaaPIeIR6yIkKgCI8Jh1QQ2TIAT52mFn3r8DIIYiPmUdVGHS3KDLHhdkfMU_KatK57ktHa_eiBebipbOVEW5GdD31pH3zcpLy82m5p4-drTJiJvql54tnpGvymznz9mJNStPF4fZY_PbyWx8H0yf7h7Go2mQSSUxUDZHGdkhYqqUiNLmK4GGUBiANDEmzmxOCgWRECpOrUyzVCplIMpjGeUke-ym9Wau9N6R1VtXrI2rNYLeJ9FNEr1P0qBXLbrdpWvK_8BDgwYIWuCrWFH9r0gvJqOD8LrlrSm1WbrC6_mLAJQAMYghKvkD7q5zpQ</recordid><startdate>200709</startdate><enddate>200709</enddate><creator>Mirisola, Mario G</creator><creator>Colomba, Leonarda</creator><creator>Gallo, Alessia</creator><creator>Amodeo, Roberta</creator><creator>De Leo, Giacomo</creator><general>John Wiley & Sons, Ltd</general><scope>FBQ</scope><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope></search><sort><creationdate>200709</creationdate><title>Yeast vectors for the integration/expression of any sequence at the TYR1 locus</title><author>Mirisola, Mario G ; Colomba, Leonarda ; Gallo, Alessia ; Amodeo, Roberta ; De Leo, Giacomo</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c3731-7fd136f411b7726b52821ae12a00b9aa8cfde712ee2278bf3bcb377a06d836de3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2007</creationdate><topic>Base Sequence</topic><topic>DNA, Fungal - chemistry</topic><topic>DNA, Fungal - genetics</topic><topic>ends‐out gene targeting</topic><topic>expression vectors</topic><topic>Gene Expression Regulation, Fungal</topic><topic>Genes, Reporter - genetics</topic><topic>Genetic Vectors - chemistry</topic><topic>Genetic Vectors - genetics</topic><topic>Green Fluorescent Proteins - genetics</topic><topic>heterologous expression</topic><topic>Molecular Sequence Data</topic><topic>Mutagenesis, Insertional - methods</topic><topic>Polymerase Chain Reaction</topic><topic>Saccharomyces cerevisiae - genetics</topic><topic>Saccharomyces cerevisiae - growth & development</topic><topic>Sequence Analysis, DNA</topic><topic>TYR1 locus</topic><topic>Tyrosine - chemistry</topic><topic>Tyrosine - genetics</topic><topic>yeast integration vector</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Mirisola, Mario G</creatorcontrib><creatorcontrib>Colomba, Leonarda</creatorcontrib><creatorcontrib>Gallo, Alessia</creatorcontrib><creatorcontrib>Amodeo, Roberta</creatorcontrib><creatorcontrib>De Leo, Giacomo</creatorcontrib><collection>AGRIS</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><jtitle>Yeast (Chichester, England)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Mirisola, Mario G</au><au>Colomba, Leonarda</au><au>Gallo, Alessia</au><au>Amodeo, Roberta</au><au>De Leo, Giacomo</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Yeast vectors for the integration/expression of any sequence at the TYR1 locus</atitle><jtitle>Yeast (Chichester, England)</jtitle><addtitle>Yeast</addtitle><date>2007-09</date><risdate>2007</risdate><volume>24</volume><issue>9</issue><spage>761</spage><epage>766</epage><pages>761-766</pages><issn>0749-503X</issn><eissn>1097-0061</eissn><abstract>We have constructed new yeast vectors for targeted integration and conditional expression of any sequence at the Saccharomyces cerevisiae TYR1 locus which becomes disrupted. We show that vector integration is not neutral, causing prototrophy for tyrosine and auxotrophy for the vector's selectable marker (uracil or leucine, depending on the vector used). This feature allows a double screening of transformed yeast cells, improving the identification of colonies with the desired chromosomal structure. The GAL10 gene promoter has been added to drive conditional expression of cloned sequences. Using these vectors, chromosomal structure verification of recombinant clones is no longer necessary, since the noise of non-homologous recombination, as well as spontaneous reversion of the selected phenotype, can easily be identified. The ability of the vector to conditionally control gene expression has been confirmed using the gene for the green fluorescent protein (GFP) as a reporter. GenBank Accession Nos EF202083-202086. Copyright © 2007 John Wiley & Sons, Ltd.</abstract><cop>Chichester, UK</cop><pub>John Wiley & Sons, Ltd</pub><pmid>17597490</pmid><doi>10.1002/yea.1511</doi><tpages>6</tpages><oa>free_for_read</oa></addata></record> |
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| source | MEDLINE; Wiley Online Library Journals Frontfile Complete; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Wiley Free Content |
| subjects | Base Sequence DNA, Fungal - chemistry DNA, Fungal - genetics ends‐out gene targeting expression vectors Gene Expression Regulation, Fungal Genes, Reporter - genetics Genetic Vectors - chemistry Genetic Vectors - genetics Green Fluorescent Proteins - genetics heterologous expression Molecular Sequence Data Mutagenesis, Insertional - methods Polymerase Chain Reaction Saccharomyces cerevisiae - genetics Saccharomyces cerevisiae - growth & development Sequence Analysis, DNA TYR1 locus Tyrosine - chemistry Tyrosine - genetics yeast integration vector |
| title | Yeast vectors for the integration/expression of any sequence at the TYR1 locus |
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