A visible phagemid system for the estimation of Cre-mediated recombination efficiency
Diversity in phage antibody libraries is important for obtaining useful high-affinity antibodies. The Cre-lox system is generally employed to increase the size of phage antibody libraries. However, estimation of library sizes after Cre-mediated recombination is difficult, since time-consuming nucleo...
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| Veröffentlicht in: | Journal of immunological methods 2003-09, Vol.280 (1), p.165-173 |
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| container_title | Journal of immunological methods |
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| creator | Kwon, Myung-Hee Lee, Myung-Shin Hong, Seung-Ho Kim, Kyongmin Hwang Shin, Ho-Joon Park, Sun Lee, Chi-Hyung Kim, Hyung-Il |
| description | Diversity in phage antibody libraries is important for obtaining useful high-affinity antibodies. The
Cre-lox system is generally employed to increase the size of phage antibody libraries. However, estimation of library sizes after Cre-mediated recombination is difficult, since time-consuming nucleotide sequence analyses are required. We have developed a visible phagemid vector system that facilitates the estimation of recombination efficiency between V
H and V
L genes. In this system, intermolecular recombination between V
L genes flanked by incompatible
lox sites is coincident with the expression of functional β-galactosidase. Recombination efficiency can be calculated simply by counting the number of blue colonies on X-gal-containing medium. Molecular analyses of plasmids isolated from blue colonies reveal a novel V
H/V
L combination. Our results suggest that this newly developed visible phagemid system may be reliably used for the measurement of recombination efficiency, which would enable precise evaluation of the diversity of phage antibody libraries. |
| doi_str_mv | 10.1016/S0022-1759(03)00261-8 |
| format | Article |
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Cre-lox system is generally employed to increase the size of phage antibody libraries. However, estimation of library sizes after Cre-mediated recombination is difficult, since time-consuming nucleotide sequence analyses are required. We have developed a visible phagemid vector system that facilitates the estimation of recombination efficiency between V
H and V
L genes. In this system, intermolecular recombination between V
L genes flanked by incompatible
lox sites is coincident with the expression of functional β-galactosidase. Recombination efficiency can be calculated simply by counting the number of blue colonies on X-gal-containing medium. Molecular analyses of plasmids isolated from blue colonies reveal a novel V
H/V
L combination. Our results suggest that this newly developed visible phagemid system may be reliably used for the measurement of recombination efficiency, which would enable precise evaluation of the diversity of phage antibody libraries.</description><identifier>ISSN: 0022-1759</identifier><identifier>EISSN: 1872-7905</identifier><identifier>DOI: 10.1016/S0022-1759(03)00261-8</identifier><identifier>PMID: 12972197</identifier><identifier>CODEN: JIMMBG</identifier><language>eng</language><publisher>Amsterdam: Elsevier B.V</publisher><subject>Amino Acid Sequence ; Animals ; Antibodies, immunoglobulins ; Antibody library size ; Base Sequence ; Biological and medical sciences ; Cre-mediated recombination ; DNA, Recombinant - genetics ; Fundamental and applied biological sciences. Psychology ; Fundamental immunology ; Genes, Immunoglobulin ; Genetic Vectors ; Immunoglobulin Heavy Chains - genetics ; Immunoglobulin Light Chains - genetics ; Immunoglobulin Variable Region - genetics ; Integrases - genetics ; Mice ; Molecular immunology ; Monoclonal antibodies ; Peptide Library ; Recombination efficiency ; Recombination, Genetic ; Viral Proteins - genetics ; Visible phagemid system ; X-gal staining</subject><ispartof>Journal of immunological methods, 2003-09, Vol.280 (1), p.165-173</ispartof><rights>2003</rights><rights>2004 INIST-CNRS</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c422t-64cbde2e0f7c76e3f70420a4303fb6d13a56a95ccc44ea94fe28f24692abd3483</citedby><cites>FETCH-LOGICAL-c422t-64cbde2e0f7c76e3f70420a4303fb6d13a56a95ccc44ea94fe28f24692abd3483</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0022175903002618$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,776,780,3537,27901,27902,65306</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=15121946$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12972197$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Kwon, Myung-Hee</creatorcontrib><creatorcontrib>Lee, Myung-Shin</creatorcontrib><creatorcontrib>Hong, Seung-Ho</creatorcontrib><creatorcontrib>Kim, Kyongmin Hwang</creatorcontrib><creatorcontrib>Shin, Ho-Joon</creatorcontrib><creatorcontrib>Park, Sun</creatorcontrib><creatorcontrib>Lee, Chi-Hyung</creatorcontrib><creatorcontrib>Kim, Hyung-Il</creatorcontrib><title>A visible phagemid system for the estimation of Cre-mediated recombination efficiency</title><title>Journal of immunological methods</title><addtitle>J Immunol Methods</addtitle><description>Diversity in phage antibody libraries is important for obtaining useful high-affinity antibodies. The
Cre-lox system is generally employed to increase the size of phage antibody libraries. However, estimation of library sizes after Cre-mediated recombination is difficult, since time-consuming nucleotide sequence analyses are required. We have developed a visible phagemid vector system that facilitates the estimation of recombination efficiency between V
H and V
L genes. In this system, intermolecular recombination between V
L genes flanked by incompatible
lox sites is coincident with the expression of functional β-galactosidase. Recombination efficiency can be calculated simply by counting the number of blue colonies on X-gal-containing medium. Molecular analyses of plasmids isolated from blue colonies reveal a novel V
H/V
L combination. Our results suggest that this newly developed visible phagemid system may be reliably used for the measurement of recombination efficiency, which would enable precise evaluation of the diversity of phage antibody libraries.</description><subject>Amino Acid Sequence</subject><subject>Animals</subject><subject>Antibodies, immunoglobulins</subject><subject>Antibody library size</subject><subject>Base Sequence</subject><subject>Biological and medical sciences</subject><subject>Cre-mediated recombination</subject><subject>DNA, Recombinant - genetics</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Fundamental immunology</subject><subject>Genes, Immunoglobulin</subject><subject>Genetic Vectors</subject><subject>Immunoglobulin Heavy Chains - genetics</subject><subject>Immunoglobulin Light Chains - genetics</subject><subject>Immunoglobulin Variable Region - genetics</subject><subject>Integrases - genetics</subject><subject>Mice</subject><subject>Molecular immunology</subject><subject>Monoclonal antibodies</subject><subject>Peptide Library</subject><subject>Recombination efficiency</subject><subject>Recombination, Genetic</subject><subject>Viral Proteins - genetics</subject><subject>Visible phagemid system</subject><subject>X-gal staining</subject><issn>0022-1759</issn><issn>1872-7905</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2003</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkElrHDEQRkVIiMfLT0jQJSE-tF1aWhqdjBniBQw52HMWanUpo9DLROoxzL-PZiE--iQKvar66hHyhcEVA6aunwE4r5iuzQ8Ql6VQrJp_IDM217zSBuqPZPYfOSGnOf8BAAYKPpMTxo3mzOgZWd7S15hj0yFdr9xv7GNL8zZP2NMwJjqtkGKeYu-mOA50DHSRsOqxjW7Clib0Y9_E4fCLIUQfcfDbc_IpuC7jxfE9I8u7ny-Lh-rp1_3j4vap8pLzqVLSNy1yhKC9ViiCBsnBSQEiNKplwtXKmdp7LyU6IwPyeeBSGe6aVsi5OCPfD3PXafy7KUFtH7PHrnMDjptstVBKi5q9CzLDlK6lKWB9AH0ac04Y7DqV69PWMrA78XYv3u6sWhB2L97uknw9Ltg0Rc9b19F0Ab4dAZe960Jyg4_5jatZwaQq3M2Bw-LtNWKyee-0KC-2J9uO8Z0o_wA8vp-c</recordid><startdate>20030901</startdate><enddate>20030901</enddate><creator>Kwon, Myung-Hee</creator><creator>Lee, Myung-Shin</creator><creator>Hong, Seung-Ho</creator><creator>Kim, Kyongmin Hwang</creator><creator>Shin, Ho-Joon</creator><creator>Park, Sun</creator><creator>Lee, Chi-Hyung</creator><creator>Kim, Hyung-Il</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>7T5</scope><scope>H94</scope><scope>7X8</scope></search><sort><creationdate>20030901</creationdate><title>A visible phagemid system for the estimation of Cre-mediated recombination efficiency</title><author>Kwon, Myung-Hee ; Lee, Myung-Shin ; Hong, Seung-Ho ; Kim, Kyongmin Hwang ; Shin, Ho-Joon ; Park, Sun ; Lee, Chi-Hyung ; Kim, Hyung-Il</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c422t-64cbde2e0f7c76e3f70420a4303fb6d13a56a95ccc44ea94fe28f24692abd3483</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2003</creationdate><topic>Amino Acid Sequence</topic><topic>Animals</topic><topic>Antibodies, immunoglobulins</topic><topic>Antibody library size</topic><topic>Base Sequence</topic><topic>Biological and medical sciences</topic><topic>Cre-mediated recombination</topic><topic>DNA, Recombinant - genetics</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>Fundamental immunology</topic><topic>Genes, Immunoglobulin</topic><topic>Genetic Vectors</topic><topic>Immunoglobulin Heavy Chains - genetics</topic><topic>Immunoglobulin Light Chains - genetics</topic><topic>Immunoglobulin Variable Region - genetics</topic><topic>Integrases - genetics</topic><topic>Mice</topic><topic>Molecular immunology</topic><topic>Monoclonal antibodies</topic><topic>Peptide Library</topic><topic>Recombination efficiency</topic><topic>Recombination, Genetic</topic><topic>Viral Proteins - genetics</topic><topic>Visible phagemid system</topic><topic>X-gal staining</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Kwon, Myung-Hee</creatorcontrib><creatorcontrib>Lee, Myung-Shin</creatorcontrib><creatorcontrib>Hong, Seung-Ho</creatorcontrib><creatorcontrib>Kim, Kyongmin Hwang</creatorcontrib><creatorcontrib>Shin, Ho-Joon</creatorcontrib><creatorcontrib>Park, Sun</creatorcontrib><creatorcontrib>Lee, Chi-Hyung</creatorcontrib><creatorcontrib>Kim, Hyung-Il</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>Immunology Abstracts</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>MEDLINE - Academic</collection><jtitle>Journal of immunological methods</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Kwon, Myung-Hee</au><au>Lee, Myung-Shin</au><au>Hong, Seung-Ho</au><au>Kim, Kyongmin Hwang</au><au>Shin, Ho-Joon</au><au>Park, Sun</au><au>Lee, Chi-Hyung</au><au>Kim, Hyung-Il</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>A visible phagemid system for the estimation of Cre-mediated recombination efficiency</atitle><jtitle>Journal of immunological methods</jtitle><addtitle>J Immunol Methods</addtitle><date>2003-09-01</date><risdate>2003</risdate><volume>280</volume><issue>1</issue><spage>165</spage><epage>173</epage><pages>165-173</pages><issn>0022-1759</issn><eissn>1872-7905</eissn><coden>JIMMBG</coden><abstract>Diversity in phage antibody libraries is important for obtaining useful high-affinity antibodies. The
Cre-lox system is generally employed to increase the size of phage antibody libraries. However, estimation of library sizes after Cre-mediated recombination is difficult, since time-consuming nucleotide sequence analyses are required. We have developed a visible phagemid vector system that facilitates the estimation of recombination efficiency between V
H and V
L genes. In this system, intermolecular recombination between V
L genes flanked by incompatible
lox sites is coincident with the expression of functional β-galactosidase. Recombination efficiency can be calculated simply by counting the number of blue colonies on X-gal-containing medium. Molecular analyses of plasmids isolated from blue colonies reveal a novel V
H/V
L combination. Our results suggest that this newly developed visible phagemid system may be reliably used for the measurement of recombination efficiency, which would enable precise evaluation of the diversity of phage antibody libraries.</abstract><cop>Amsterdam</cop><pub>Elsevier B.V</pub><pmid>12972197</pmid><doi>10.1016/S0022-1759(03)00261-8</doi><tpages>9</tpages></addata></record> |
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| ispartof | Journal of immunological methods, 2003-09, Vol.280 (1), p.165-173 |
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| subjects | Amino Acid Sequence Animals Antibodies, immunoglobulins Antibody library size Base Sequence Biological and medical sciences Cre-mediated recombination DNA, Recombinant - genetics Fundamental and applied biological sciences. Psychology Fundamental immunology Genes, Immunoglobulin Genetic Vectors Immunoglobulin Heavy Chains - genetics Immunoglobulin Light Chains - genetics Immunoglobulin Variable Region - genetics Integrases - genetics Mice Molecular immunology Monoclonal antibodies Peptide Library Recombination efficiency Recombination, Genetic Viral Proteins - genetics Visible phagemid system X-gal staining |
| title | A visible phagemid system for the estimation of Cre-mediated recombination efficiency |
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