Enzymatic amplification of latent pseudorabies virus nucleic acid sequences
Identifieur interne : 001033 ( Istex/Corpus ); précédent : 001032; suivant : 001034Enzymatic amplification of latent pseudorabies virus nucleic acid sequences
Auteurs : J. R Lokensgard ; D. G. Thawley ; T. W. MolitorSource :
- Journal of Virological Methods [ 0166-0934 ] ; 1991.
English descriptors
- Teeft :
- Acute phase, Amplification, Amplification techniques, Antisense orientation, Assay, Bamhl fragment, Buffer control, Cells harbor, Cheung, Coding strand, Denatured ssdna, Denhardt solution, Enzymatic amplification, Ethidium bromide, Ganglion, Gene expression, Genome, Hybridization, Internal oligonucleotide, Internal probe, Latency, Latent pseudorabies virus, Latent state, Latently, Nucleic, Nucleic acid amplification methods, Nucleic acids, Oligonucleotide, Polymerase, Polymerase chain reaction, Positive control, Primer, Pseudorabies, Pseudorabies virus, Pseudorabies virus genome, Room temperature, Saint paul, Same polarity, Sequence data, Sodium phosphate, Standard dntps, Swine, Template strand, Trigeminal ganglia, Uninfected swine, Vero cells, Viral, Viral gene expression, Virol.
Abstract
Abstract: To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. These nucleic acid amplification methods should be applicable to the investigation of PRV latency, and gene expression during latency and reactivation, in which few cells harbor latent virus.
Url:
DOI: 10.1016/0166-0934(91)90120-O
Links to Exploration step
ISTEX:1D6488862892600F56DB36C7AEB7DC76FD838D0ALe document en format XML
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R Lokensgard</name><affiliation><mods:affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</mods:affiliation></affiliation></author><author><name sortKey="Thawley, D G" sort="Thawley, D G" uniqKey="Thawley D" first="D. G." last="Thawley">D. G. Thawley</name><affiliation><mods:affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</mods:affiliation></affiliation></author><author><name sortKey="Molitor, T W" sort="Molitor, T W" uniqKey="Molitor T" first="T. W." last="Molitor">T. W. Molitor</name><affiliation><mods:affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</mods:affiliation></affiliation></author></analytic><monogr></monogr><series><title level="j">Journal of Virological Methods</title><title level="j" type="abbrev">VIRMET</title><idno type="ISSN">0166-0934</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="1991">1991</date><biblScope unit="volume">34</biblScope><biblScope unit="issue">1</biblScope><biblScope unit="page" from="45">45</biblScope><biblScope unit="page" to="55">55</biblScope></imprint><idno type="ISSN">0166-0934</idno></series></biblStruct></sourceDesc><seriesStmt><idno type="ISSN">0166-0934</idno></seriesStmt></fileDesc><profileDesc><textClass><keywords scheme="Teeft" xml:lang="en"><term>Acute phase</term><term>Amplification</term><term>Amplification techniques</term><term>Antisense orientation</term><term>Assay</term><term>Bamhl fragment</term><term>Buffer control</term><term>Cells harbor</term><term>Cheung</term><term>Coding strand</term><term>Denatured ssdna</term><term>Denhardt solution</term><term>Enzymatic amplification</term><term>Ethidium bromide</term><term>Ganglion</term><term>Gene expression</term><term>Genome</term><term>Hybridization</term><term>Internal oligonucleotide</term><term>Internal probe</term><term>Latency</term><term>Latent pseudorabies virus</term><term>Latent state</term><term>Latently</term><term>Nucleic</term><term>Nucleic acid amplification methods</term><term>Nucleic acids</term><term>Oligonucleotide</term><term>Polymerase</term><term>Polymerase chain reaction</term><term>Positive control</term><term>Primer</term><term>Pseudorabies</term><term>Pseudorabies virus</term><term>Pseudorabies virus genome</term><term>Room temperature</term><term>Saint paul</term><term>Same polarity</term><term>Sequence data</term><term>Sodium phosphate</term><term>Standard dntps</term><term>Swine</term><term>Template strand</term><term>Trigeminal ganglia</term><term>Uninfected swine</term><term>Vero cells</term><term>Viral</term><term>Viral gene expression</term><term>Virol</term></keywords></textClass><langUsage><language ident="en">en</language></langUsage></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Abstract: To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. 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Molitor</name><affiliations><json:string>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</json:string></affiliations></json:item></author><subject><json:item><lang><json:string>eng</json:string></lang><value>Pseudorabies virus</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Latency</value></json:item><json:item><lang><json:string>eng</json:string></lang><value>Polymerase chain reaction</value></json:item></subject><arkIstex>ark:/67375/6H6-WPWQB8GC-R</arkIstex><language><json:string>eng</json:string></language><originalGenre><json:string>Full-length article</json:string></originalGenre><abstract>Abstract: To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. These nucleic acid amplification methods should be applicable to the investigation of PRV latency, and gene expression during latency and reactivation, in which few cells harbor latent virus.</abstract><qualityIndicators><score>7.795</score><pdfWordCount>3515</pdfWordCount><pdfCharCount>21161</pdfCharCount><pdfVersion>1.4</pdfVersion><pdfPageCount>11</pdfPageCount><pdfPageSize>432 x 612 pts</pdfPageSize><refBibsNative>true</refBibsNative><abstractWordCount>190</abstractWordCount><abstractCharCount>1283</abstractCharCount><keywordCount>3</keywordCount></qualityIndicators><title>Enzymatic amplification of latent pseudorabies virus nucleic acid sequences</title><pmid><json:string>1715897</json:string></pmid><pii><json:string>0166-0934(91)90120-O</json:string></pii><genre><json:string>research-article</json:string></genre><serie><title>Proc. Natl. Acad. Sci. 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Fig</json:string><json:string>A. Primer</json:string><json:string>D.G. Thawley</json:string><json:string>Previous</json:string><json:string>C. Primer</json:string><json:string>In</json:string><json:string>U.S.A. Sciences</json:string><json:string>T.W. Molitor</json:string><json:string>La Jolla</json:string><json:string>C. 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Molitor, Dept. of Clinical and Population Sciences, 225 Veterinary Teaching Hospitals, 1365 Gortner Avenue, Saint Paul, MN 55108, U.S.A.</p></note><affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</affiliation></author><idno type="istex">1D6488862892600F56DB36C7AEB7DC76FD838D0A</idno><idno type="ark">ark:/67375/6H6-WPWQB8GC-R</idno><idno type="DOI">10.1016/0166-0934(91)90120-O</idno><idno type="PII">0166-0934(91)90120-O</idno></analytic><monogr><title level="j">Journal of Virological Methods</title><title level="j" type="abbrev">VIRMET</title><idno type="pISSN">0166-0934</idno><idno type="PII">S0166-0934(00)X0207-8</idno><imprint><publisher>ELSEVIER</publisher><date type="published" when="1991"></date><biblScope unit="volume">34</biblScope><biblScope unit="issue">1</biblScope><biblScope unit="page" from="45">45</biblScope><biblScope unit="page" to="55">55</biblScope></imprint></monogr></biblStruct></sourceDesc></fileDesc><profileDesc><creation><date>1991</date></creation><langUsage><language ident="en">en</language></langUsage><abstract xml:lang="en"><p>Abstract: To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. These nucleic acid amplification methods should be applicable to the investigation of PRV latency, and gene expression during latency and reactivation, in which few cells harbor latent virus.</p></abstract><textClass><keywords scheme="keyword"><list><head>Keywords</head><item><term>Pseudorabies virus</term></item><item><term>Latency</term></item><item><term>Polymerase chain reaction</term></item></list></keywords></textClass></profileDesc><revisionDesc><change when="1991">Published</change></revisionDesc></teiHeader></istex:fulltextTEI><json:item><extension>txt</extension><original>false</original><mimetype>text/plain</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPWQB8GC-R/fulltext.txt</uri></json:item></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:docType><istex:document><converted-article version="4.5.2" docsubtype="fla"><item-info><jid>VIRMET</jid><aid>9190120O</aid><ce:pii>0166-0934(91)90120-O</ce:pii><ce:doi>10.1016/0166-0934(91)90120-O</ce:doi><ce:copyright type="unknown" year="1991"></ce:copyright></item-info><head><ce:title>Enzymatic amplification of latent pseudorabies virus nucleic acid sequences</ce:title><ce:author-group><ce:author><ce:given-name>J.R</ce:given-name><ce:surname>Lokensgard</ce:surname></ce:author><ce:author><ce:given-name>D.G.</ce:given-name><ce:surname>Thawley</ce:surname></ce:author><ce:author><ce:given-name>T.W.</ce:given-name><ce:surname>Molitor</ce:surname><ce:cross-ref refid="COR1"><ce:sup>∗</ce:sup></ce:cross-ref></ce:author><ce:affiliation><ce:textfn>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</ce:textfn></ce:affiliation><ce:correspondence id="COR1"><ce:label>∗</ce:label><ce:text>Correspondence to: Dr. T.W. Molitor, Dept. of Clinical and Population Sciences, 225 Veterinary Teaching Hospitals, 1365 Gortner Avenue, Saint Paul, MN 55108, U.S.A.</ce:text></ce:correspondence></ce:author-group><ce:date-accepted day="9" month="4" year="1991"></ce:date-accepted><ce:abstract><ce:section-title>Abstract</ce:section-title><ce:abstract-sec><ce:simple-para>To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. These nucleic acid amplification methods should be applicable to the investigation of PRV latency, and gene expression during latency and reactivation, in which few cells harbor latent virus.</ce:simple-para></ce:abstract-sec></ce:abstract><ce:keywords><ce:section-title>Keywords</ce:section-title><ce:keyword><ce:text>Pseudorabies virus</ce:text></ce:keyword><ce:keyword><ce:text>Latency</ce:text></ce:keyword><ce:keyword><ce:text>Polymerase chain reaction</ce:text></ce:keyword></ce:keywords></head></converted-article></istex:document></istex:metadataXml><mods version="3.6"><titleInfo><title>Enzymatic amplification of latent pseudorabies virus nucleic acid sequences</title></titleInfo><titleInfo type="alternative" contentType="CDATA"><title>Enzymatic amplification of latent pseudorabies virus nucleic acid sequences</title></titleInfo><name type="personal"><namePart type="given">J.R</namePart><namePart type="family">Lokensgard</namePart><affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">D.G.</namePart><namePart type="family">Thawley</namePart><affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</affiliation><role><roleTerm type="text">author</roleTerm></role></name><name type="personal"><namePart type="given">T.W.</namePart><namePart type="family">Molitor</namePart><affiliation>Department of Clinical and Population Sciences, College of Veterinary Medicine, University of Minnesota, Saint Paul, Minnesota, U.S.A.</affiliation><description>Correspondence to: Dr. T.W. Molitor, Dept. of Clinical and Population Sciences, 225 Veterinary Teaching Hospitals, 1365 Gortner Avenue, Saint Paul, MN 55108, U.S.A.</description><role><roleTerm type="text">author</roleTerm></role></name><typeOfResource>text</typeOfResource><genre type="research-article" displayLabel="Full-length article" authority="ISTEX" authorityURI="https://content-type.data.istex.fr" valueURI="https://content-type.data.istex.fr/ark:/67375/XTP-1JC4F85T-7">research-article</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">1991</dateIssued><copyrightDate encoding="w3cdtf">1991</copyrightDate></originInfo><language><languageTerm type="code" authority="iso639-2b">eng</languageTerm><languageTerm type="code" authority="rfc3066">en</languageTerm></language><abstract lang="en">Abstract: To investigate various aspects of the latency of pseudorabies virus in swine (PRV, suid herpesvirus 1) we developed in vitro nucleic acid amplification methods based upon the polymerase chain reaction. Primers flanking a 156-bp region of the pseudorabies virus gp II gene were annealed to purified PRV DNA as well as DNA isolated from the trigeminal ganglia of swine latently infected with PRV and subjected to PCR amplification. Following amplification, 100 fg of PRV DNA was visualizable on stained gels and 1 fg (equivalent to 6 viral genome copies) was detectable when amplification was combined with blot hybridization. PRV-specific DNA sequences which remained undetectable by direct blot hybridization assays were amplified to levels visualizable on ethidium-bromide-stained gels in 5 of 5 experimental latently infected animals. In addition, oligonucleotide primers specific for a 223-bp region of the PRV immediate-early gene (IE 180) were capable of amplifying overlapping latency associated transcripts (LATs), via a cDNA intermediate, in 6 of 6 latently infected swine. These nucleic acid amplification methods should be applicable to the investigation of PRV latency, and gene expression during latency and reactivation, in which few cells harbor latent virus.</abstract><subject><genre>Keywords</genre><topic>Pseudorabies virus</topic><topic>Latency</topic><topic>Polymerase chain reaction</topic></subject><relatedItem type="host"><titleInfo><title>Journal of Virological Methods</title></titleInfo><titleInfo type="abbreviated"><title>VIRMET</title></titleInfo><genre type="journal" authority="ISTEX" authorityURI="https://publication-type.data.istex.fr" valueURI="https://publication-type.data.istex.fr/ark:/67375/JMC-0GLKJH51-B">journal</genre><originInfo><publisher>ELSEVIER</publisher><dateIssued encoding="w3cdtf">1991</dateIssued></originInfo><identifier type="ISSN">0166-0934</identifier><identifier type="PII">S0166-0934(00)X0207-8</identifier><part><date>1991</date><detail type="volume"><number>34</number><caption>vol.</caption></detail><detail type="issue"><number>1</number><caption>no.</caption></detail><extent unit="issue-pages"><start>1</start><end>112</end></extent><extent unit="pages"><start>45</start><end>55</end></extent></part></relatedItem><identifier type="istex">1D6488862892600F56DB36C7AEB7DC76FD838D0A</identifier><identifier type="ark">ark:/67375/6H6-WPWQB8GC-R</identifier><identifier type="DOI">10.1016/0166-0934(91)90120-O</identifier><identifier type="PII">0166-0934(91)90120-O</identifier><recordInfo><recordContentSource authority="ISTEX" authorityURI="https://loaded-corpus.data.istex.fr" valueURI="https://loaded-corpus.data.istex.fr/ark:/67375/XBH-HKKZVM7B-M">elsevier</recordContentSource></recordInfo></mods><json:item><extension>json</extension><original>false</original><mimetype>application/json</mimetype><uri>https://api.istex.fr/ark:/67375/6H6-WPWQB8GC-R/record.json</uri></json:item></metadata></istex></record>Pour manipuler ce document sous Unix (Dilib)
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