Loading…
Regulated gene expression in transfected primary chicken erythrocytes
We describe a method for studying transient gene expression in primary avian erythroid cells that involves controlled osmotic shock, followed by DNA transfection using DEAE-dextran. Cells treated in this way reproducibly express high levels of chloramphenicol acetyltransferase (CAT) when transfected...
Saved in:
| Published in: | Proceedings of the National Academy of Sciences - PNAS 1986-06, Vol.83 (12), p.4312-4316 |
|---|---|
| Main Authors: | , , , |
| Format: | Article |
| Language: | English |
| Subjects: | |
| Citations: | Items that cite this one |
| Online Access: | Get full text |
| Tags: |
Add Tag
No Tags, Be the first to tag this record!
|
| cited_by | cdi_FETCH-LOGICAL-c509t-4a105a7f590fbe7e2daae91f354ef945a77d0efc2542164e6c684fdee9381eca3 |
|---|---|
| cites | |
| container_end_page | 4316 |
| container_issue | 12 |
| container_start_page | 4312 |
| container_title | Proceedings of the National Academy of Sciences - PNAS |
| container_volume | 83 |
| creator | Hesse, J.E Nickol, J.M Lieber, M.R Felsenfeld, G |
| description | We describe a method for studying transient gene expression in primary avian erythroid cells that involves controlled osmotic shock, followed by DNA transfection using DEAE-dextran. Cells treated in this way reproducibly express high levels of chloramphenicol acetyltransferase (CAT) when transfected with a plasmid having the cat gene coupled to an appropriate viral promoter. An observed correlation between levels of CAT expression and extent of hemoglobin release during controlled shock makes it possible to choose optimum conditions for expression in erythroid cells at various stages of embryonic development. Using these techniques, we have investigated the effect on CAT expression of fusing to the cat gene various portions of the chicken adult (β -globin (β A) gene. We show that in 9-day or 12-day embryonic erythrocytes, the promoter activity of the 5′ flanking region of the β A gene (in the absence of any viral promoters) is strongly stimulated by a downstream sequence, located in the region 110-588 base pairs on the 3′ side of the poly(A) signal, that acts as an enhancer. Its activity is reduced in 5-day embryonic cells and absent in primary chicken fibroblasts and mouse L cells, suggesting that this transient expression system will be useful in studying developmentally regulated globin gene expression. |
| doi_str_mv | 10.1073/pnas.83.12.4312 |
| format | article |
| fullrecord | <record><control><sourceid>jstor_pasca</sourceid><recordid>TN_cdi_pascalfrancis_primary_8778211</recordid><sourceformat>XML</sourceformat><sourcesystem>PC</sourcesystem><jstor_id>27968</jstor_id><sourcerecordid>27968</sourcerecordid><originalsourceid>FETCH-LOGICAL-c509t-4a105a7f590fbe7e2daae91f354ef945a77d0efc2542164e6c684fdee9381eca3</originalsourceid><addsrcrecordid>eNp9kEtv1DAURi0EKtPCGgkJlAWCVaZ-xvaCBarKQ6qEBHRtuc71TEomntoO6vz7OpoolA0rL865937-EHpF8Jpgyc73g01rxdaErjkj9AlaEaxJ3XCNn6IVxlTWilP-HJ2mdIsx1kLhE3TCuNBEihW6_AGbsbcZ2moDA1Rwv4-QUheGqhuqHO2QPLgJ72O3s_FQuW3nfsNQQTzkbQzukCG9QM-87RO8nN8zdP358tfF1_rq-5dvF5-uaiewzjW3BAsrvdDY34AE2loLmngmOHjNC5ItBu-o4JQ0HBrXKO5bAM0UAWfZGfp43Lsfb3bQOhhKwt7M0UywnfmXDN3WbMIfwyiTlJb59_N8DHcjpGx2XXLQ93aAMCYjGyUxFU0Rz4-iiyGlCH65QbCZijdT8UYxQ6iZii8Tbx5HW_y56cLfzdwmZ3tfmnVdWjQlpaKEFO3trE37F_r4zof_CsaPfZ_hPhfz9dG8TTnERaVSN-ov9DYYu4klyvXP8nmsiGIPYL23ig</addsrcrecordid><sourcetype>Open Access Repository</sourcetype><iscdi>true</iscdi><recordtype>article</recordtype><pqid>76870256</pqid></control><display><type>article</type><title>Regulated gene expression in transfected primary chicken erythrocytes</title><source>Open Access: PubMed Central</source><source>JSTOR Archival Journals and Primary Sources Collection</source><creator>Hesse, J.E ; Nickol, J.M ; Lieber, M.R ; Felsenfeld, G</creator><creatorcontrib>Hesse, J.E ; Nickol, J.M ; Lieber, M.R ; Felsenfeld, G</creatorcontrib><description>We describe a method for studying transient gene expression in primary avian erythroid cells that involves controlled osmotic shock, followed by DNA transfection using DEAE-dextran. Cells treated in this way reproducibly express high levels of chloramphenicol acetyltransferase (CAT) when transfected with a plasmid having the cat gene coupled to an appropriate viral promoter. An observed correlation between levels of CAT expression and extent of hemoglobin release during controlled shock makes it possible to choose optimum conditions for expression in erythroid cells at various stages of embryonic development. Using these techniques, we have investigated the effect on CAT expression of fusing to the cat gene various portions of the chicken adult (β -globin (β A) gene. We show that in 9-day or 12-day embryonic erythrocytes, the promoter activity of the 5′ flanking region of the β A gene (in the absence of any viral promoters) is strongly stimulated by a downstream sequence, located in the region 110-588 base pairs on the 3′ side of the poly(A) signal, that acts as an enhancer. Its activity is reduced in 5-day embryonic cells and absent in primary chicken fibroblasts and mouse L cells, suggesting that this transient expression system will be useful in studying developmentally regulated globin gene expression.</description><identifier>ISSN: 0027-8424</identifier><identifier>EISSN: 1091-6490</identifier><identifier>DOI: 10.1073/pnas.83.12.4312</identifier><identifier>PMID: 3459175</identifier><identifier>CODEN: PNASA6</identifier><language>eng</language><publisher>Washington, DC: National Academy of Sciences of the United States of America</publisher><subject>Acetyltransferases - genetics ; Age Factors ; Ammonium Chloride - pharmacology ; Animals ; Biological and medical sciences ; Cell lines ; Cell Membrane Permeability ; Cells ; CHICKENS ; Chloramphenicol O-Acetyltransferase ; Cloning, Molecular ; CONTROL GENETICO ; DESARROLLO EMBRIONARIO ; DEVELOPPEMENT EMBRYONNAIRE ; EMBRYONIC DEVELOPMENT ; Enhancer Elements, Genetic ; ERITROCITOS ; ERYTHROCYTE ; ERYTHROCYTES ; Erythrocytes - physiology ; Erythroid cells ; Fundamental and applied biological sciences. Psychology ; GENE ; Gene expression ; Gene Expression Regulation ; GENES ; Genes, Regulator ; GENETIC CONTROL ; Genetic Vectors ; Globins - genetics ; L cells ; LUTTE GENETIQUE ; Molecular and cellular biology ; Molecular genetics ; Plasmids ; POLLO ; POULET ; Promoter regions ; Transfection</subject><ispartof>Proceedings of the National Academy of Sciences - PNAS, 1986-06, Vol.83 (12), p.4312-4316</ispartof><rights>1986 INIST-CNRS</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c509t-4a105a7f590fbe7e2daae91f354ef945a77d0efc2542164e6c684fdee9381eca3</citedby></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Uhttp://www.pnas.org/content/83/12.cover.gif</thumbnail><linktopdf>$$Uhttps://www.jstor.org/stable/pdf/27968$$EPDF$$P50$$Gjstor$$H</linktopdf><linktohtml>$$Uhttps://www.jstor.org/stable/27968$$EHTML$$P50$$Gjstor$$H</linktohtml><link.rule.ids>230,314,727,780,784,885,27924,27925,53791,53793,58238,58471</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=8778211$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/3459175$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Hesse, J.E</creatorcontrib><creatorcontrib>Nickol, J.M</creatorcontrib><creatorcontrib>Lieber, M.R</creatorcontrib><creatorcontrib>Felsenfeld, G</creatorcontrib><title>Regulated gene expression in transfected primary chicken erythrocytes</title><title>Proceedings of the National Academy of Sciences - PNAS</title><addtitle>Proc Natl Acad Sci U S A</addtitle><description>We describe a method for studying transient gene expression in primary avian erythroid cells that involves controlled osmotic shock, followed by DNA transfection using DEAE-dextran. Cells treated in this way reproducibly express high levels of chloramphenicol acetyltransferase (CAT) when transfected with a plasmid having the cat gene coupled to an appropriate viral promoter. An observed correlation between levels of CAT expression and extent of hemoglobin release during controlled shock makes it possible to choose optimum conditions for expression in erythroid cells at various stages of embryonic development. Using these techniques, we have investigated the effect on CAT expression of fusing to the cat gene various portions of the chicken adult (β -globin (β A) gene. We show that in 9-day or 12-day embryonic erythrocytes, the promoter activity of the 5′ flanking region of the β A gene (in the absence of any viral promoters) is strongly stimulated by a downstream sequence, located in the region 110-588 base pairs on the 3′ side of the poly(A) signal, that acts as an enhancer. Its activity is reduced in 5-day embryonic cells and absent in primary chicken fibroblasts and mouse L cells, suggesting that this transient expression system will be useful in studying developmentally regulated globin gene expression.</description><subject>Acetyltransferases - genetics</subject><subject>Age Factors</subject><subject>Ammonium Chloride - pharmacology</subject><subject>Animals</subject><subject>Biological and medical sciences</subject><subject>Cell lines</subject><subject>Cell Membrane Permeability</subject><subject>Cells</subject><subject>CHICKENS</subject><subject>Chloramphenicol O-Acetyltransferase</subject><subject>Cloning, Molecular</subject><subject>CONTROL GENETICO</subject><subject>DESARROLLO EMBRIONARIO</subject><subject>DEVELOPPEMENT EMBRYONNAIRE</subject><subject>EMBRYONIC DEVELOPMENT</subject><subject>Enhancer Elements, Genetic</subject><subject>ERITROCITOS</subject><subject>ERYTHROCYTE</subject><subject>ERYTHROCYTES</subject><subject>Erythrocytes - physiology</subject><subject>Erythroid cells</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>GENE</subject><subject>Gene expression</subject><subject>Gene Expression Regulation</subject><subject>GENES</subject><subject>Genes, Regulator</subject><subject>GENETIC CONTROL</subject><subject>Genetic Vectors</subject><subject>Globins - genetics</subject><subject>L cells</subject><subject>LUTTE GENETIQUE</subject><subject>Molecular and cellular biology</subject><subject>Molecular genetics</subject><subject>Plasmids</subject><subject>POLLO</subject><subject>POULET</subject><subject>Promoter regions</subject><subject>Transfection</subject><issn>0027-8424</issn><issn>1091-6490</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1986</creationdate><recordtype>article</recordtype><recordid>eNp9kEtv1DAURi0EKtPCGgkJlAWCVaZ-xvaCBarKQ6qEBHRtuc71TEomntoO6vz7OpoolA0rL865937-EHpF8Jpgyc73g01rxdaErjkj9AlaEaxJ3XCNn6IVxlTWilP-HJ2mdIsx1kLhE3TCuNBEihW6_AGbsbcZ2moDA1Rwv4-QUheGqhuqHO2QPLgJ72O3s_FQuW3nfsNQQTzkbQzukCG9QM-87RO8nN8zdP358tfF1_rq-5dvF5-uaiewzjW3BAsrvdDY34AE2loLmngmOHjNC5ItBu-o4JQ0HBrXKO5bAM0UAWfZGfp43Lsfb3bQOhhKwt7M0UywnfmXDN3WbMIfwyiTlJb59_N8DHcjpGx2XXLQ93aAMCYjGyUxFU0Rz4-iiyGlCH65QbCZijdT8UYxQ6iZii8Tbx5HW_y56cLfzdwmZ3tfmnVdWjQlpaKEFO3trE37F_r4zof_CsaPfZ_hPhfz9dG8TTnERaVSN-ov9DYYu4klyvXP8nmsiGIPYL23ig</recordid><startdate>19860601</startdate><enddate>19860601</enddate><creator>Hesse, J.E</creator><creator>Nickol, J.M</creator><creator>Lieber, M.R</creator><creator>Felsenfeld, G</creator><general>National Academy of Sciences of the United States of America</general><general>National Acad Sciences</general><scope>FBQ</scope><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>7X8</scope><scope>5PM</scope></search><sort><creationdate>19860601</creationdate><title>Regulated gene expression in transfected primary chicken erythrocytes</title><author>Hesse, J.E ; Nickol, J.M ; Lieber, M.R ; Felsenfeld, G</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c509t-4a105a7f590fbe7e2daae91f354ef945a77d0efc2542164e6c684fdee9381eca3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1986</creationdate><topic>Acetyltransferases - genetics</topic><topic>Age Factors</topic><topic>Ammonium Chloride - pharmacology</topic><topic>Animals</topic><topic>Biological and medical sciences</topic><topic>Cell lines</topic><topic>Cell Membrane Permeability</topic><topic>Cells</topic><topic>CHICKENS</topic><topic>Chloramphenicol O-Acetyltransferase</topic><topic>Cloning, Molecular</topic><topic>CONTROL GENETICO</topic><topic>DESARROLLO EMBRIONARIO</topic><topic>DEVELOPPEMENT EMBRYONNAIRE</topic><topic>EMBRYONIC DEVELOPMENT</topic><topic>Enhancer Elements, Genetic</topic><topic>ERITROCITOS</topic><topic>ERYTHROCYTE</topic><topic>ERYTHROCYTES</topic><topic>Erythrocytes - physiology</topic><topic>Erythroid cells</topic><topic>Fundamental and applied biological sciences. Psychology</topic><topic>GENE</topic><topic>Gene expression</topic><topic>Gene Expression Regulation</topic><topic>GENES</topic><topic>Genes, Regulator</topic><topic>GENETIC CONTROL</topic><topic>Genetic Vectors</topic><topic>Globins - genetics</topic><topic>L cells</topic><topic>LUTTE GENETIQUE</topic><topic>Molecular and cellular biology</topic><topic>Molecular genetics</topic><topic>Plasmids</topic><topic>POLLO</topic><topic>POULET</topic><topic>Promoter regions</topic><topic>Transfection</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Hesse, J.E</creatorcontrib><creatorcontrib>Nickol, J.M</creatorcontrib><creatorcontrib>Lieber, M.R</creatorcontrib><creatorcontrib>Felsenfeld, G</creatorcontrib><collection>AGRIS</collection><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>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Hesse, J.E</au><au>Nickol, J.M</au><au>Lieber, M.R</au><au>Felsenfeld, G</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Regulated gene expression in transfected primary chicken erythrocytes</atitle><jtitle>Proceedings of the National Academy of Sciences - PNAS</jtitle><addtitle>Proc Natl Acad Sci U S A</addtitle><date>1986-06-01</date><risdate>1986</risdate><volume>83</volume><issue>12</issue><spage>4312</spage><epage>4316</epage><pages>4312-4316</pages><issn>0027-8424</issn><eissn>1091-6490</eissn><coden>PNASA6</coden><abstract>We describe a method for studying transient gene expression in primary avian erythroid cells that involves controlled osmotic shock, followed by DNA transfection using DEAE-dextran. Cells treated in this way reproducibly express high levels of chloramphenicol acetyltransferase (CAT) when transfected with a plasmid having the cat gene coupled to an appropriate viral promoter. An observed correlation between levels of CAT expression and extent of hemoglobin release during controlled shock makes it possible to choose optimum conditions for expression in erythroid cells at various stages of embryonic development. Using these techniques, we have investigated the effect on CAT expression of fusing to the cat gene various portions of the chicken adult (β -globin (β A) gene. We show that in 9-day or 12-day embryonic erythrocytes, the promoter activity of the 5′ flanking region of the β A gene (in the absence of any viral promoters) is strongly stimulated by a downstream sequence, located in the region 110-588 base pairs on the 3′ side of the poly(A) signal, that acts as an enhancer. Its activity is reduced in 5-day embryonic cells and absent in primary chicken fibroblasts and mouse L cells, suggesting that this transient expression system will be useful in studying developmentally regulated globin gene expression.</abstract><cop>Washington, DC</cop><pub>National Academy of Sciences of the United States of America</pub><pmid>3459175</pmid><doi>10.1073/pnas.83.12.4312</doi><tpages>5</tpages><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
| identifier | ISSN: 0027-8424 |
| ispartof | Proceedings of the National Academy of Sciences - PNAS, 1986-06, Vol.83 (12), p.4312-4316 |
| issn | 0027-8424 1091-6490 |
| language | eng |
| recordid | cdi_pascalfrancis_primary_8778211 |
| source | Open Access: PubMed Central; JSTOR Archival Journals and Primary Sources Collection |
| subjects | Acetyltransferases - genetics Age Factors Ammonium Chloride - pharmacology Animals Biological and medical sciences Cell lines Cell Membrane Permeability Cells CHICKENS Chloramphenicol O-Acetyltransferase Cloning, Molecular CONTROL GENETICO DESARROLLO EMBRIONARIO DEVELOPPEMENT EMBRYONNAIRE EMBRYONIC DEVELOPMENT Enhancer Elements, Genetic ERITROCITOS ERYTHROCYTE ERYTHROCYTES Erythrocytes - physiology Erythroid cells Fundamental and applied biological sciences. Psychology GENE Gene expression Gene Expression Regulation GENES Genes, Regulator GENETIC CONTROL Genetic Vectors Globins - genetics L cells LUTTE GENETIQUE Molecular and cellular biology Molecular genetics Plasmids POLLO POULET Promoter regions Transfection |
| title | Regulated gene expression in transfected primary chicken erythrocytes |
| url | http://sfxeu10.hosted.exlibrisgroup.com/loughborough?ctx_ver=Z39.88-2004&ctx_enc=info:ofi/enc:UTF-8&ctx_tim=2025-01-04T03%3A54%3A25IST&url_ver=Z39.88-2004&url_ctx_fmt=infofi/fmt:kev:mtx:ctx&rfr_id=info:sid/primo.exlibrisgroup.com:primo3-Article-jstor_pasca&rft_val_fmt=info:ofi/fmt:kev:mtx:journal&rft.genre=article&rft.atitle=Regulated%20gene%20expression%20in%20transfected%20primary%20chicken%20erythrocytes&rft.jtitle=Proceedings%20of%20the%20National%20Academy%20of%20Sciences%20-%20PNAS&rft.au=Hesse,%20J.E&rft.date=1986-06-01&rft.volume=83&rft.issue=12&rft.spage=4312&rft.epage=4316&rft.pages=4312-4316&rft.issn=0027-8424&rft.eissn=1091-6490&rft.coden=PNASA6&rft_id=info:doi/10.1073/pnas.83.12.4312&rft_dat=%3Cjstor_pasca%3E27968%3C/jstor_pasca%3E%3Cgrp_id%3Ecdi_FETCH-LOGICAL-c509t-4a105a7f590fbe7e2daae91f354ef945a77d0efc2542164e6c684fdee9381eca3%3C/grp_id%3E%3Coa%3E%3C/oa%3E%3Curl%3E%3C/url%3E&rft_id=info:oai/&rft_pqid=76870256&rft_id=info:pmid/3459175&rft_jstor_id=27968&rfr_iscdi=true |