Use of bacterial DHFR-II fusion proteins to elicit specific antibodies
Plasmids containing the coding region of the type II dihydrofolate reductase (DHFR) specified by R388 have been used to alter the amino acid (aa) sequence at the C-terminus of this protein. These plasmids have a unique cloning site in the C-terminal portion of the 78-aa coding region. Insertions of...
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| Veröffentlicht in: | Gene 1986, Vol.41 (2), p.289-297 |
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| creator | Vermersch, P.S. Klass, M.R. Benne, G.N. |
| description | Plasmids containing the coding region of the type II dihydrofolate reductase (DHFR) specified by R388 have been used to alter the amino acid (aa) sequence at the C-terminus of this protein. These plasmids have a unique cloning site in the C-terminal portion of the 78-aa coding region. Insertions of DNA fragments into this site produced plasmids that code for proteins with 6- to 80-aa extensions. The vectors were constructed to terminate translation in all three phases beyond the position of insertion of foreign DNA.
Random DNA fragments from the major sperm protein (MSP) gene of
Caenorhabditis elegans produced by DNase I cleavage were inserted into these vectors. Cell extracts from colonies containing MSP sequences were examined by gel electrophoresis and immunoblotting. One of the hybrid DHFR-MSP proteins was isolated and antibody was prepared to it. This antibody preparation reacted with MSP in immunoblots of purified MSP and whole cell extracts of the worm. A rapid purification procedure for the DHFR is presented. |
| doi_str_mv | 10.1016/0378-1119(86)90109-5 |
| format | Article |
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Random DNA fragments from the major sperm protein (MSP) gene of
Caenorhabditis elegans produced by DNase I cleavage were inserted into these vectors. Cell extracts from colonies containing MSP sequences were examined by gel electrophoresis and immunoblotting. One of the hybrid DHFR-MSP proteins was isolated and antibody was prepared to it. This antibody preparation reacted with MSP in immunoblots of purified MSP and whole cell extracts of the worm. A rapid purification procedure for the DHFR is presented.</description><identifier>ISSN: 0378-1119</identifier><identifier>EISSN: 1879-0038</identifier><identifier>DOI: 10.1016/0378-1119(86)90109-5</identifier><identifier>PMID: 3011601</identifier><identifier>CODEN: GENED6</identifier><language>eng</language><publisher>Lausanne: Elsevier B.V</publisher><subject>Amino Acid Sequence ; Antibodies ; Antigen-Antibody Complex ; Biological and medical sciences ; Biotechnology ; Caenorhabditis elegans ; Caenorhabditis elegans, trimethoprim resistance ; DNA Restriction Enzymes ; Escherichia coli - enzymology ; Escherichia coli - genetics ; Fundamental and applied biological sciences. Psychology ; Genetic engineering ; Genetic technics ; Genetic Vectors ; major sperm protein ; Methods. Procedures. Technologies ; Molecular cloning ; Nucleic Acid Hybridization ; plasmid vectors ; Plasmids ; Recombinant DNA ; Tetrahydrofolate Dehydrogenase - genetics ; Tetrahydrofolate Dehydrogenase - immunology ; translation</subject><ispartof>Gene, 1986, Vol.41 (2), p.289-297</ispartof><rights>1986</rights><rights>1987 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c417t-92f835f237159e3288bb70e874139ece596981a9fa1c66f43e53362fd9d732543</citedby><cites>FETCH-LOGICAL-c417t-92f835f237159e3288bb70e874139ece596981a9fa1c66f43e53362fd9d732543</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/0378-1119(86)90109-5$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,780,784,3548,4022,27922,27923,27924,45994</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=7956520$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/3011601$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Vermersch, P.S.</creatorcontrib><creatorcontrib>Klass, M.R.</creatorcontrib><creatorcontrib>Benne, G.N.</creatorcontrib><title>Use of bacterial DHFR-II fusion proteins to elicit specific antibodies</title><title>Gene</title><addtitle>Gene</addtitle><description>Plasmids containing the coding region of the type II dihydrofolate reductase (DHFR) specified by R388 have been used to alter the amino acid (aa) sequence at the C-terminus of this protein. These plasmids have a unique cloning site in the C-terminal portion of the 78-aa coding region. Insertions of DNA fragments into this site produced plasmids that code for proteins with 6- to 80-aa extensions. The vectors were constructed to terminate translation in all three phases beyond the position of insertion of foreign DNA.
Random DNA fragments from the major sperm protein (MSP) gene of
Caenorhabditis elegans produced by DNase I cleavage were inserted into these vectors. Cell extracts from colonies containing MSP sequences were examined by gel electrophoresis and immunoblotting. One of the hybrid DHFR-MSP proteins was isolated and antibody was prepared to it. This antibody preparation reacted with MSP in immunoblots of purified MSP and whole cell extracts of the worm. A rapid purification procedure for the DHFR is presented.</description><subject>Amino Acid Sequence</subject><subject>Antibodies</subject><subject>Antigen-Antibody Complex</subject><subject>Biological and medical sciences</subject><subject>Biotechnology</subject><subject>Caenorhabditis elegans</subject><subject>Caenorhabditis elegans, trimethoprim resistance</subject><subject>DNA Restriction Enzymes</subject><subject>Escherichia coli - enzymology</subject><subject>Escherichia coli - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Genetic engineering</subject><subject>Genetic technics</subject><subject>Genetic Vectors</subject><subject>major sperm protein</subject><subject>Methods. Procedures. Technologies</subject><subject>Molecular cloning</subject><subject>Nucleic Acid Hybridization</subject><subject>plasmid vectors</subject><subject>Plasmids</subject><subject>Recombinant DNA</subject><subject>Tetrahydrofolate Dehydrogenase - genetics</subject><subject>Tetrahydrofolate Dehydrogenase - immunology</subject><subject>translation</subject><issn>0378-1119</issn><issn>1879-0038</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1986</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkE1rGzEQhkVJSN2k_6AFHUJpD9to9K1LoKR1YggEQnMWWu0IVNa7jrQO5N93XRsf07nMYZ6ZeXkI-QTsOzDQV0wY2wCA-2r1N8eAuUa9IwuwxjWMCXtCFkfkPflQ6x82l1L8jJwJBqAZLMjyqSIdE21DnLDk0NOfd8vHZrWiaVvzONBNGSfMQ6XTSLHPMU-0bjDmlCMNw5TbsctYL8hpCn3Fj4d-Tp6Wv37f3DX3D7ermx_3TZRgpsbxZIVKXBhQDgW3tm0NQ2skCIcRldPOQnApQNQ6SYFKCM1T5zojuJLinHzZ351jPW-xTn6da8S-DwOO2-qNttJKx_8LglRacMlmUO7BWMZaCya_KXkdyqsH5nee_U6i30n0Vvt_nr2a1z4f7m_bNXbHpYPYeX55mIcaQ59KGGKuR8w4pRXffb_eYzhLe8lYfI0Zh4hdLhgn34357Rx_Ad1Plk8</recordid><startdate>1986</startdate><enddate>1986</enddate><creator>Vermersch, P.S.</creator><creator>Klass, M.R.</creator><creator>Benne, G.N.</creator><general>Elsevier B.V</general><general>Elsevier</general><scope>IQODW</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><scope>7TM</scope><scope>8FD</scope><scope>FR3</scope><scope>P64</scope><scope>RC3</scope><scope>7X8</scope></search><sort><creationdate>1986</creationdate><title>Use of bacterial DHFR-II fusion proteins to elicit specific antibodies</title><author>Vermersch, P.S. ; Klass, M.R. ; Benne, G.N.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c417t-92f835f237159e3288bb70e874139ece596981a9fa1c66f43e53362fd9d732543</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1986</creationdate><topic>Amino Acid Sequence</topic><topic>Antibodies</topic><topic>Antigen-Antibody Complex</topic><topic>Biological and medical sciences</topic><topic>Biotechnology</topic><topic>Caenorhabditis elegans</topic><topic>Caenorhabditis elegans, trimethoprim resistance</topic><topic>DNA Restriction Enzymes</topic><topic>Escherichia coli - enzymology</topic><topic>Escherichia coli - genetics</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Genetic engineering</topic><topic>Genetic technics</topic><topic>Genetic Vectors</topic><topic>major sperm protein</topic><topic>Methods. Procedures. Technologies</topic><topic>Molecular cloning</topic><topic>Nucleic Acid Hybridization</topic><topic>plasmid vectors</topic><topic>Plasmids</topic><topic>Recombinant DNA</topic><topic>Tetrahydrofolate Dehydrogenase - genetics</topic><topic>Tetrahydrofolate Dehydrogenase - immunology</topic><topic>translation</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Vermersch, P.S.</creatorcontrib><creatorcontrib>Klass, M.R.</creatorcontrib><creatorcontrib>Benne, G.N.</creatorcontrib><collection>Pascal-Francis</collection><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><collection>Technology Research Database</collection><collection>Engineering Research Database</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Gene</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Vermersch, P.S.</au><au>Klass, M.R.</au><au>Benne, G.N.</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Use of bacterial DHFR-II fusion proteins to elicit specific antibodies</atitle><jtitle>Gene</jtitle><addtitle>Gene</addtitle><date>1986</date><risdate>1986</risdate><volume>41</volume><issue>2</issue><spage>289</spage><epage>297</epage><pages>289-297</pages><issn>0378-1119</issn><eissn>1879-0038</eissn><coden>GENED6</coden><abstract>Plasmids containing the coding region of the type II dihydrofolate reductase (DHFR) specified by R388 have been used to alter the amino acid (aa) sequence at the C-terminus of this protein. These plasmids have a unique cloning site in the C-terminal portion of the 78-aa coding region. Insertions of DNA fragments into this site produced plasmids that code for proteins with 6- to 80-aa extensions. The vectors were constructed to terminate translation in all three phases beyond the position of insertion of foreign DNA.
Random DNA fragments from the major sperm protein (MSP) gene of
Caenorhabditis elegans produced by DNase I cleavage were inserted into these vectors. Cell extracts from colonies containing MSP sequences were examined by gel electrophoresis and immunoblotting. One of the hybrid DHFR-MSP proteins was isolated and antibody was prepared to it. This antibody preparation reacted with MSP in immunoblots of purified MSP and whole cell extracts of the worm. A rapid purification procedure for the DHFR is presented.</abstract><cop>Lausanne</cop><cop>Amsterdam</cop><cop>New York, NY</cop><pub>Elsevier B.V</pub><pmid>3011601</pmid><doi>10.1016/0378-1119(86)90109-5</doi><tpages>9</tpages></addata></record> |
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| identifier | ISSN: 0378-1119 |
| ispartof | Gene, 1986, Vol.41 (2), p.289-297 |
| issn | 0378-1119 1879-0038 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_76848492 |
| source | MEDLINE; ScienceDirect Journals (5 years ago - present) |
| subjects | Amino Acid Sequence Antibodies Antigen-Antibody Complex Biological and medical sciences Biotechnology Caenorhabditis elegans Caenorhabditis elegans, trimethoprim resistance DNA Restriction Enzymes Escherichia coli - enzymology Escherichia coli - genetics Fundamental and applied biological sciences. Psychology Genetic engineering Genetic technics Genetic Vectors major sperm protein Methods. Procedures. Technologies Molecular cloning Nucleic Acid Hybridization plasmid vectors Plasmids Recombinant DNA Tetrahydrofolate Dehydrogenase - genetics Tetrahydrofolate Dehydrogenase - immunology translation |
| title | Use of bacterial DHFR-II fusion proteins to elicit specific antibodies |
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