Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325
Identifieur interne : 001499 ( Istex/Corpus ); précédent : 001498; suivant : 001500Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325
Auteurs : Joan L. Betz ; John R. SadlerSource :
- Gene [ 0378-1119 ] ; 1981.
Abstract
The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the EcoRI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cmr revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: In the revertant pOE223, the deletion converts the “upstream” EcoRI site to GAATC (a site for HinfI), while in pOE222 both EcoRI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cmr phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cms phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: Spontaneous, as well as mutT-induced, Cmr revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.
Url:
DOI: 10.1016/0378-1119(81)90128-1
Links to Exploration step
ISTEX:5D7739B809C836E48F65130F14230C0EA56F6F26Le document en format XML
<record><TEI wicri:istexFullTextTei="biblStruct"><teiHeader><fileDesc><titleStmt><title>Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
<author><name sortKey="Betz, Joan L" sort="Betz, Joan L" uniqKey="Betz J" first="Joan L." last="Betz">Joan L. Betz</name>
<affiliation><mods:affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</mods:affiliation>
</affiliation>
</author>
<author><name sortKey="Sadler, John R" sort="Sadler, John R" uniqKey="Sadler J" first="John R." last="Sadler">John R. Sadler</name>
<affiliation><mods:affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</mods:affiliation>
</affiliation>
</author>
</titleStmt>
<publicationStmt><idno type="wicri:source">ISTEX</idno>
<idno type="RBID">ISTEX:5D7739B809C836E48F65130F14230C0EA56F6F26</idno>
<date when="1981" year="1981">1981</date>
<idno type="doi">10.1016/0378-1119(81)90128-1</idno>
<idno type="url">https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/fulltext/pdf</idno>
<idno type="wicri:Area/Istex/Corpus">001499</idno>
</publicationStmt>
<sourceDesc><biblStruct><analytic><title level="a">Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
<author><name sortKey="Betz, Joan L" sort="Betz, Joan L" uniqKey="Betz J" first="Joan L." last="Betz">Joan L. Betz</name>
<affiliation><mods:affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</mods:affiliation>
</affiliation>
</author>
<author><name sortKey="Sadler, John R" sort="Sadler, John R" uniqKey="Sadler J" first="John R." last="Sadler">John R. Sadler</name>
<affiliation><mods:affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</mods:affiliation>
</affiliation>
</author>
</analytic>
<monogr></monogr>
<series><title level="j">Gene</title>
<title level="j" type="abbrev">GENE</title>
<idno type="ISSN">0378-1119</idno>
<imprint><publisher>ELSEVIER</publisher>
<date type="published" when="1981">1981</date>
<biblScope unit="volume">15</biblScope>
<biblScope unit="issue">2–3</biblScope>
<biblScope unit="page" from="187">187</biblScope>
<biblScope unit="page" to="200">200</biblScope>
</imprint>
<idno type="ISSN">0378-1119</idno>
</series>
<idno type="istex">5D7739B809C836E48F65130F14230C0EA56F6F26</idno>
<idno type="DOI">10.1016/0378-1119(81)90128-1</idno>
<idno type="PII">0378-1119(81)90128-1</idno>
</biblStruct>
</sourceDesc>
<seriesStmt><idno type="ISSN">0378-1119</idno>
</seriesStmt>
</fileDesc>
<profileDesc><textClass></textClass>
<langUsage><language ident="en">en</language>
</langUsage>
</profileDesc>
</teiHeader>
<front><div type="abstract" xml:lang="en">The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the EcoRI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cmr revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: In the revertant pOE223, the deletion converts the “upstream” EcoRI site to GAATC (a site for HinfI), while in pOE222 both EcoRI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cmr phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cms phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: Spontaneous, as well as mutT-induced, Cmr revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.</div>
</front>
</TEI>
<istex><corpusName>elsevier</corpusName>
<author><json:item><name>Joan L. Betz</name>
<affiliations><json:string>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</json:string>
</affiliations>
</json:item>
<json:item><name>John R. Sadler</name>
<affiliations><json:string>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</json:string>
</affiliations>
</json:item>
</author>
<subject><json:item><lang><json:string>eng</json:string>
</lang>
<value>Recombinant DNA</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>frameshift mutations</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>chain-terminating codons</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>operator-encoded peptide segment</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>BAP : bacterial alkaline phosphatase</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>bp : base pair(s)</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>CAT : chloramphenicol transacetylase</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>Cm : chloramphenicol</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>C.T. : chain-terminating</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>IPTG : isopropylthio-β-d-galactose</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>Tc : tetracycline</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>661 buffer : 6 mM Tris pH 7.5 at 25°C, 6 mM NaCl, 1 mM EDTA</value>
</json:item>
<json:item><lang><json:string>eng</json:string>
</lang>
<value>[] : indicates plasmid-carrier state</value>
</json:item>
</subject>
<language><json:string>eng</json:string>
</language>
<abstract>The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the EcoRI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cmr revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: In the revertant pOE223, the deletion converts the “upstream” EcoRI site to GAATC (a site for HinfI), while in pOE222 both EcoRI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cmr phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cms phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: Spontaneous, as well as mutT-induced, Cmr revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.</abstract>
<qualityIndicators><score>7.808</score>
<pdfVersion>1.3</pdfVersion>
<pdfPageSize>576 x 792 pts</pdfPageSize>
<refBibsNative>true</refBibsNative>
<keywordCount>13</keywordCount>
<abstractCharCount>1480</abstractCharCount>
<pdfWordCount>5826</pdfWordCount>
<pdfCharCount>35214</pdfCharCount>
<pdfPageCount>14</pdfPageCount>
<abstractWordCount>234</abstractWordCount>
</qualityIndicators>
<title>Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
<pii><json:string>0378-1119(81)90128-1</json:string>
</pii>
<genre><json:string>research-article</json:string>
</genre>
<serie><volume>75</volume>
<pages><last>3582</last>
<first>3578</first>
</pages>
<genre></genre>
<language><json:string>unknown</json:string>
</language>
<title>Proc. Natl. Acad. Sci. USA</title>
</serie>
<host><volume>15</volume>
<pii><json:string>S0378-1119(00)X0479-9</json:string>
</pii>
<pages><last>200</last>
<first>187</first>
</pages>
<issn><json:string>0378-1119</json:string>
</issn>
<issue>2–3</issue>
<genre><json:string>Journal</json:string>
</genre>
<language><json:string>unknown</json:string>
</language>
<title>Gene</title>
<publicationDate>1981</publicationDate>
</host>
<categories><wos><json:string>BIOCHEMISTRY & MOLECULAR BIOLOGY</json:string>
<json:string>GENETICS & HEREDITY</json:string>
</wos>
</categories>
<publicationDate>1981</publicationDate>
<copyrightDate>1981</copyrightDate>
<doi><json:string>10.1016/0378-1119(81)90128-1</json:string>
</doi>
<id>5D7739B809C836E48F65130F14230C0EA56F6F26</id>
<fulltext><json:item><original>true</original>
<mimetype>application/pdf</mimetype>
<extension>pdf</extension>
<uri>https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/fulltext/pdf</uri>
</json:item>
<json:item><original>true</original>
<mimetype>text/plain</mimetype>
<extension>txt</extension>
<uri>https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/fulltext/txt</uri>
</json:item>
<json:item><original>false</original>
<mimetype>application/zip</mimetype>
<extension>zip</extension>
<uri>https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/fulltext/zip</uri>
</json:item>
<istex:fulltextTEI uri="https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/fulltext/tei"><teiHeader><fileDesc><titleStmt><title level="a">Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
</titleStmt>
<publicationStmt><authority>ISTEX</authority>
<publisher>ELSEVIER</publisher>
<availability><p>ELSEVIER</p>
</availability>
<date>1981</date>
</publicationStmt>
<notesStmt><note>Communicated by H.O. Smith</note>
</notesStmt>
<sourceDesc><biblStruct type="inbook"><analytic><title level="a">Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
<author><persName><forename type="first">Joan L.</forename>
<surname>Betz</surname>
</persName>
<note type="biography">To whom reprint requests should be addressed.</note>
<affiliation>To whom reprint requests should be addressed.</affiliation>
<affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</affiliation>
</author>
<author><persName><forename type="first">John R.</forename>
<surname>Sadler</surname>
</persName>
<affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</affiliation>
</author>
</analytic>
<monogr><title level="j">Gene</title>
<title level="j" type="abbrev">GENE</title>
<idno type="pISSN">0378-1119</idno>
<idno type="PII">S0378-1119(00)X0479-9</idno>
<imprint><publisher>ELSEVIER</publisher>
<date type="published" when="1981"></date>
<biblScope unit="volume">15</biblScope>
<biblScope unit="issue">2–3</biblScope>
<biblScope unit="page" from="187">187</biblScope>
<biblScope unit="page" to="200">200</biblScope>
</imprint>
</monogr>
<idno type="istex">5D7739B809C836E48F65130F14230C0EA56F6F26</idno>
<idno type="DOI">10.1016/0378-1119(81)90128-1</idno>
<idno type="PII">0378-1119(81)90128-1</idno>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc><creation><date>1981</date>
</creation>
<langUsage><language ident="en">en</language>
</langUsage>
<abstract xml:lang="en"><p>The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the EcoRI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cmr revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: In the revertant pOE223, the deletion converts the “upstream” EcoRI site to GAATC (a site for HinfI), while in pOE222 both EcoRI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cmr phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cms phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: Spontaneous, as well as mutT-induced, Cmr revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.</p>
</abstract>
<textClass><keywords scheme="keyword"><list><head>Keywords</head>
<item><term>Recombinant DNA</term>
</item>
<item><term>frameshift mutations</term>
</item>
<item><term>chain-terminating codons</term>
</item>
<item><term>operator-encoded peptide segment</term>
</item>
</list>
</keywords>
</textClass>
<textClass><keywords scheme="keyword"><list><head>Abbreviations</head>
<item><term>BAP</term>
<term>bacterial alkaline phosphatase</term>
</item>
<item><term>bp</term>
<term>base pair(s)</term>
</item>
<item><term>CAT</term>
<term>chloramphenicol transacetylase</term>
</item>
<item><term>Cm</term>
<term>chloramphenicol</term>
</item>
<item><term>C.T.</term>
<term>chain-terminating</term>
</item>
<item><term>IPTG</term>
<term>isopropylthio-β-d-galactose</term>
</item>
<item><term>Tc</term>
<term>tetracycline</term>
</item>
<item><term>661 buffer</term>
<term>6 mM Tris pH 7.5 at 25°C, 6 mM NaCl, 1 mM EDTA</term>
</item>
<item><term>[]</term>
<term>indicates plasmid-carrier state</term>
</item>
</list>
</keywords>
</textClass>
</profileDesc>
<revisionDesc><change when="1981-06-08">Registration</change>
<change when="1981">Published</change>
</revisionDesc>
</teiHeader>
</istex:fulltextTEI>
</fulltext>
<metadata><istex:metadataXml wicri:clean="Elsevier, elements deleted: tail"><istex:xmlDeclaration>version="1.0" encoding="UTF-8" </istex:xmlDeclaration>
<istex:docType PUBLIC="-//ES//DTD journal article DTD version 4.5.2//EN//XML" URI="art452.dtd" name="istex:docType"><istex:entity SYSTEM="fx1" NDATA="IMAGE" name="fx1"></istex:entity>
<istex:entity SYSTEM="fx2" NDATA="IMAGE" name="fx2"></istex:entity>
</istex:docType>
<istex:document><converted-article version="4.5.2" docsubtype="fla"><item-info><jid>GENE</jid>
<aid>90128</aid>
<ce:pii>0378-1119(81)90128-1</ce:pii>
<ce:doi>10.1016/0378-1119(81)90128-1</ce:doi>
<ce:copyright type="unknown" year="1981"></ce:copyright>
</item-info>
<head><ce:title>Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</ce:title>
<ce:author-group><ce:author><ce:given-name>Joan L.</ce:given-name>
<ce:surname>Betz</ce:surname>
<ce:cross-ref refid="COR1"><ce:sup>∗</ce:sup>
</ce:cross-ref>
</ce:author>
<ce:author><ce:given-name>John R.</ce:given-name>
<ce:surname>Sadler</ce:surname>
</ce:author>
<ce:affiliation><ce:textfn>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</ce:textfn>
</ce:affiliation>
<ce:correspondence id="COR1"><ce:label>∗</ce:label>
<ce:text>To whom reprint requests should be addressed.</ce:text>
</ce:correspondence>
</ce:author-group>
<ce:date-received day="15" month="5" year="1981"></ce:date-received>
<ce:date-accepted day="8" month="6" year="1981"></ce:date-accepted>
<ce:miscellaneous>Communicated by H.O. Smith</ce:miscellaneous>
<ce:abstract><ce:section-title>Abstract</ce:section-title>
<ce:abstract-sec><ce:simple-para id="SP0005">The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the <ce:italic>Eco</ce:italic>
RI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cm<ce:sup>r</ce:sup>
revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: <ce:inline-figure><ce:link locator="fx1"></ce:link>
</ce:inline-figure>
</ce:simple-para>
<ce:simple-para id="SP0010">In the revertant pOE223, the deletion converts the “upstream” <ce:italic>Eco</ce:italic>
RI site to GAATC (a site for <ce:italic>Hin</ce:italic>
fI), while in pOE222 both <ce:italic>Eco</ce:italic>
RI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cm<ce:sup>r</ce:sup>
phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cm<ce:sup>s</ce:sup>
phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: <ce:inline-figure><ce:link locator="fx2"></ce:link>
</ce:inline-figure>
</ce:simple-para>
<ce:simple-para id="SP0015">Spontaneous, as well as <ce:italic>mutT</ce:italic>
-induced, Cm<ce:sup>r</ce:sup>
revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.</ce:simple-para>
</ce:abstract-sec>
</ce:abstract>
<ce:keywords><ce:section-title>Keywords</ce:section-title>
<ce:keyword><ce:text>Recombinant DNA</ce:text>
</ce:keyword>
<ce:keyword><ce:text>frameshift mutations</ce:text>
</ce:keyword>
<ce:keyword><ce:text>chain-terminating codons</ce:text>
</ce:keyword>
<ce:keyword><ce:text>operator-encoded peptide segment</ce:text>
</ce:keyword>
</ce:keywords>
<ce:keywords class="abr"><ce:section-title>Abbreviations</ce:section-title>
<ce:keyword><ce:text>BAP</ce:text>
<ce:keyword><ce:text>bacterial alkaline phosphatase</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>bp</ce:text>
<ce:keyword><ce:text>base pair(s)</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>CAT</ce:text>
<ce:keyword><ce:text>chloramphenicol transacetylase</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>Cm</ce:text>
<ce:keyword><ce:text>chloramphenicol</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>C.T.</ce:text>
<ce:keyword><ce:text>chain-terminating</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>IPTG</ce:text>
<ce:keyword><ce:text>isopropylthio-β-<ce:small-caps>d</ce:small-caps>
-galactose</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>Tc</ce:text>
<ce:keyword><ce:text>tetracycline</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>661 buffer</ce:text>
<ce:keyword><ce:text>6 mM Tris pH 7.5 at 25°C, 6 mM NaCl, 1 mM EDTA</ce:text>
</ce:keyword>
</ce:keyword>
<ce:keyword><ce:text>[]</ce:text>
<ce:keyword><ce:text>indicates plasmid-carrier state</ce:text>
</ce:keyword>
</ce:keyword>
</ce:keywords>
</head>
</converted-article>
</istex:document>
</istex:metadataXml>
<mods version="3.6"><titleInfo><title>Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
</titleInfo>
<titleInfo type="alternative" contentType="CDATA"><title>Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325</title>
</titleInfo>
<name type="personal"><namePart type="given">Joan L.</namePart>
<namePart type="family">Betz</namePart>
<affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</affiliation>
<description>To whom reprint requests should be addressed.</description>
<role><roleTerm type="text">author</roleTerm>
</role>
</name>
<name type="personal"><namePart type="given">John R.</namePart>
<namePart type="family">Sadler</namePart>
<affiliation>Department of Biochemistry, Biophysics and Genetics, University of Colorado Health Sciences Center, Denver, CO 80262, U.S.A.</affiliation>
<role><roleTerm type="text">author</roleTerm>
</role>
</name>
<typeOfResource>text</typeOfResource>
<genre type="research-article" displayLabel="Full-length article"></genre>
<originInfo><publisher>ELSEVIER</publisher>
<dateIssued encoding="w3cdtf">1981</dateIssued>
<dateValid encoding="w3cdtf">1981-06-08</dateValid>
<copyrightDate encoding="w3cdtf">1981</copyrightDate>
</originInfo>
<language><languageTerm type="code" authority="iso639-2b">eng</languageTerm>
<languageTerm type="code" authority="rfc3066">en</languageTerm>
</language>
<physicalDescription><internetMediaType>text/html</internetMediaType>
</physicalDescription>
<abstract lang="en">The chloramphenicol transacetylase (CAT) gene of plasmid pBR325 is inactivated by insertion of a 40-bp lactose operator DNA fragment into the EcoRI site located 216 bp downstream of the initiation codon of the gene. Rare spontaneous Cmr revertants have been selected, some of which still contain a functional operator in the CAT gene. DNA sequencing in the operator region of two revertant plasmids revealed that both had suffered single base-pair deletions just upstream of the operator segment: In the revertant pOE223, the deletion converts the “upstream” EcoRI site to GAATC (a site for HinfI), while in pOE222 both EcoRI sites flanking the operator remain intact. Removal of the operator from pOE222 creates a new plasmid pJT7 in which CAT gene contains a (−1) frameshift at position 213. Reinsertion of the 40-bp operator, in the orientation shown above, recreates pOE222 and restores the Cmr phenotype. However, insertion of the operator in the reverse orientation into pJT7creates a new plasmid, pOE271, with a Cms phenotype due to an in-phase TAA chain terminating (C.T.) codon in the operator as shown: Spontaneous, as well as mutT-induced, Cmr revertants of pOE271 have been recovered which still contain a 40-bp operator; ten have been sequenced and shown to contain single-base substitutions at positions 14 or 15 in the operator sequence in the C.T. codon. Three of these mutations significantly reduce repressor affinity for the operator, while the fourth does not.</abstract>
<note>Communicated by H.O. Smith</note>
<subject><genre>Keywords</genre>
<topic>Recombinant DNA</topic>
<topic>frameshift mutations</topic>
<topic>chain-terminating codons</topic>
<topic>operator-encoded peptide segment</topic>
</subject>
<subject><genre>Abbreviations</genre>
<topic>BAP : bacterial alkaline phosphatase</topic>
<topic>bp : base pair(s)</topic>
<topic>CAT : chloramphenicol transacetylase</topic>
<topic>Cm : chloramphenicol</topic>
<topic>C.T. : chain-terminating</topic>
<topic>IPTG : isopropylthio-β-d-galactose</topic>
<topic>Tc : tetracycline</topic>
<topic>661 buffer : 6 mM Tris pH 7.5 at 25°C, 6 mM NaCl, 1 mM EDTA</topic>
<topic>[] : indicates plasmid-carrier state</topic>
</subject>
<relatedItem type="host"><titleInfo><title>Gene</title>
</titleInfo>
<titleInfo type="abbreviated"><title>GENE</title>
</titleInfo>
<genre type="Journal">journal</genre>
<originInfo><dateIssued encoding="w3cdtf">198111</dateIssued>
</originInfo>
<identifier type="ISSN">0378-1119</identifier>
<identifier type="PII">S0378-1119(00)X0479-9</identifier>
<part><date>198111</date>
<detail type="volume"><number>15</number>
<caption>vol.</caption>
</detail>
<detail type="issue"><number>2–3</number>
<caption>no.</caption>
</detail>
<extent unit="issue pages"><start>103</start>
<end>296</end>
</extent>
<extent unit="pages"><start>187</start>
<end>200</end>
</extent>
</part>
</relatedItem>
<identifier type="istex">5D7739B809C836E48F65130F14230C0EA56F6F26</identifier>
<identifier type="DOI">10.1016/0378-1119(81)90128-1</identifier>
<identifier type="PII">0378-1119(81)90128-1</identifier>
<recordInfo><recordContentSource>ELSEVIER</recordContentSource>
</recordInfo>
</mods>
</metadata>
<enrichments><istex:catWosTEI uri="https://api.istex.fr/document/5D7739B809C836E48F65130F14230C0EA56F6F26/enrichments/catWos"><teiHeader><profileDesc><textClass><classCode scheme="WOS">BIOCHEMISTRY & MOLECULAR BIOLOGY</classCode>
<classCode scheme="WOS">GENETICS & HEREDITY</classCode>
</textClass>
</profileDesc>
</teiHeader>
</istex:catWosTEI>
</enrichments>
</istex>
</record>
Pour manipuler ce document sous Unix (Dilib)
EXPLOR_STEP=$WICRI_ROOT/Wicri/Musique/explor/OperaV1/Data/Istex/Corpus
HfdSelect -h $EXPLOR_STEP/biblio.hfd -nk 001499 | SxmlIndent | more
Ou
HfdSelect -h $EXPLOR_AREA/Data/Istex/Corpus/biblio.hfd -nk 001499 | SxmlIndent | more
Pour mettre un lien sur cette page dans le réseau Wicri
{{Explor lien |wiki= Wicri/Musique |area= OperaV1 |flux= Istex |étape= Corpus |type= RBID |clé= ISTEX:5D7739B809C836E48F65130F14230C0EA56F6F26 |texte= Variants of a cloned synthetic lactose operator II. Chloramphenicol-resistant revertants retaining a lactose operator in the CAT gene of plasmid pBR325 }}
![]() | This area was generated with Dilib version V0.6.21. | ![]() |