SrasV1, Main, Exploration, bibRecord, 005172

Real-Time Quantitative Fluorescent Reverse Transcriptase-PCR for Detection of Severe Acute Respiratory Syndrome-Associated Coronavirus RNA

Identifieur interne : 005172 ( Main/Exploration ); précédent : 005171; suivant : 005173

Real-Time Quantitative Fluorescent Reverse Transcriptase-PCR for Detection of Severe Acute Respiratory Syndrome-Associated Coronavirus RNA

Source :

Molecular Diagnosis [ 1084-8592 ] ; 2004.

RBID : PMC:7100218

Descripteurs français

KwdFr :
- ARN viral (analyse), ARN viral (génétique), Amorces ADN, Fluorescence, Humains, RT-PCR (), Sensibilité et spécificité, Syndrome respiratoire aigu sévère (virologie), Valeurs de référence, Virus du SRAS (génétique), Virus du SRAS (isolement et purification), Études cas-témoins.
MESH :
- analyse : ARN viral.
- génétique : ARN viral, Virus du SRAS.
- isolement et purification : Virus du SRAS.
- virologie : Syndrome respiratoire aigu sévère.
- Amorces ADN, Fluorescence, Humains, RT-PCR, Sensibilité et spécificité, Valeurs de référence, Études cas-témoins.

English descriptors

KwdEn :
- Case-Control Studies, DNA Primers, Fluorescence, Humans, RNA, Viral (analysis), RNA, Viral (genetics), Reference Values, Reverse Transcriptase Polymerase Chain Reaction (methods), SARS Virus (genetics), SARS Virus (isolation & purification), Sensitivity and Specificity, Severe Acute Respiratory Syndrome (virology).
MESH :
- chemical , analysis : RNA, Viral.
- chemical , genetics : RNA, Viral.
- chemical : DNA Primers.
- genetics : SARS Virus.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction.
- virology : Severe Acute Respiratory Syndrome.
- Case-Control Studies, Fluorescence, Humans, Reference Values, Sensitivity and Specificity.

Abstract

Aim: SARS-associated coronavirus (SARS-CoV) has been confirmed as the pathogen for severe acute respiratory syndrome (SARS). The aim of our study was to construct a sensitive and specific real-time quantitative fluorescent (QF) reverse transcriptase (RT)-PCR method for the detection of SARS-CoV RNA.

Methods: Stored blood specimens from 44 patients with confirmed SARS were used along with blood samples from two sets of controls, 30 healthy volunteers who had no contact with SARS patients, and 30 healthy doctors and nurses who had contact with SARS patients but were without symptoms of SARS. Two pairs of primers were synthesized by the Shanghai Sangon Company according to SARS-CoV BJ01 strain sequence (AY278488), and then a pair of primers were designed and compared with a pair of primers published by WHO.

Results: Using serial dilutions of SARS-CoV, the 44 blood samples from SARS patients specimens were tested. Using a 0.01% dilution of SARS-CoV, all 44 clinical samples tested positive in our assay. In comparison, using a 0.1% dilution of SARS-CoV, 26 of the 44 samples tested positive using the WHO primers. In the QF-RT-PCR assay, there was a linear amplification from 100 copies to 10⁸ copies of the control RNA per RT-PCR and at least 10 copies, and sometimes even 1 copy, of target RNA tested positive in our assay.

Conclusion: The primer we developed is sufficiently sensitive and specific to diagnose symptomatic SARS-CoV infections and for monitoring virus load.

Url:

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7100218

DOI: 10.1007/BF03260067
PubMed: 15887978
PubMed Central: 7100218

Affiliations:

Le document en format XML

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<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Gan, Haixue" sort="Gan, Haixue" uniqKey="Gan H" first="Haixue" last="Gan">Haixue Gan</name>
<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Huang, Shengyong" sort="Huang, Shengyong" uniqKey="Huang S" first="Shengyong" last="Huang">Shengyong Huang</name>
<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Wen, Jie" sort="Wen, Jie" uniqKey="Wen J" first="Jie" last="Wen">Jie Wen</name>
<affiliation wicri:level="3"><nlm:aff id="Aff1"><institution-wrap><institution-id institution-id-type="GRID">grid.9227.e</institution-id>
<institution-id institution-id-type="ISNI">0000000119573309</institution-id>
<institution>Beijing Genomics Institute,</institution>
<institution>Chinese Academy of Sciences,</institution>
</institution-wrap>
Beijing Airport Industry Zone B-6, Beijing, 101300 China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing Airport Industry Zone B-6, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Fang, Jianqiu" sort="Fang, Jianqiu" uniqKey="Fang J" first="Jianqiu" last="Fang">Jianqiu Fang</name>
<affiliation wicri:level="3"><nlm:aff id="Aff1"><institution-wrap><institution-id institution-id-type="GRID">grid.9227.e</institution-id>
<institution-id institution-id-type="ISNI">0000000119573309</institution-id>
<institution>Beijing Genomics Institute,</institution>
<institution>Chinese Academy of Sciences,</institution>
</institution-wrap>
Beijing Airport Industry Zone B-6, Beijing, 101300 China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing Airport Industry Zone B-6, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
<author><name sortKey="Wang, Jian" sort="Wang, Jian" uniqKey="Wang J" first="Jian" last="Wang">Jian Wang</name>
<affiliation wicri:level="3"><nlm:aff id="Aff1"><institution-wrap><institution-id institution-id-type="GRID">grid.9227.e</institution-id>
<institution-id institution-id-type="ISNI">0000000119573309</institution-id>
<institution>Beijing Genomics Institute,</institution>
<institution>Chinese Academy of Sciences,</institution>
</institution-wrap>
Beijing Airport Industry Zone B-6, Beijing, 101300 China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing Airport Industry Zone B-6, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
<affiliation wicri:level="3"><nlm:aff id="Aff2">Beijing BGI-GBI Biotech Co. Ltd, Beijing, China</nlm:aff>
<country xml:lang="fr">République populaire de Chine</country>
<wicri:regionArea>Beijing BGI-GBI Biotech Co. Ltd, Beijing</wicri:regionArea>
<placeName><settlement type="city">Pékin</settlement>
</placeName>
</affiliation>
</author>
</analytic>
<series><title level="j">Molecular Diagnosis</title>
<idno type="ISSN">1084-8592</idno>
<idno type="eISSN">1179-2000</idno>
<imprint><date when="2004">2004</date>
</imprint>
</series>
</biblStruct>
</sourceDesc>
</fileDesc>
<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Case-Control Studies</term>
<term>DNA Primers</term>
<term>Fluorescence</term>
<term>Humans</term>
<term>RNA, Viral (analysis)</term>
<term>RNA, Viral (genetics)</term>
<term>Reference Values</term>
<term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term>
<term>SARS Virus (genetics)</term>
<term>SARS Virus (isolation & purification)</term>
<term>Sensitivity and Specificity</term>
<term>Severe Acute Respiratory Syndrome (virology)</term>
</keywords>
<keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral (analyse)</term>
<term>ARN viral (génétique)</term>
<term>Amorces ADN</term>
<term>Fluorescence</term>
<term>Humains</term>
<term>RT-PCR ()</term>
<term>Sensibilité et spécificité</term>
<term>Syndrome respiratoire aigu sévère (virologie)</term>
<term>Valeurs de référence</term>
<term>Virus du SRAS (génétique)</term>
<term>Virus du SRAS (isolement et purification)</term>
<term>Études cas-témoins</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="analysis" xml:lang="en"><term>RNA, Viral</term>
</keywords>
<keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>RNA, Viral</term>
</keywords>
<keywords scheme="MESH" type="chemical" xml:lang="en"><term>DNA Primers</term>
</keywords>
<keywords scheme="MESH" qualifier="analyse" xml:lang="fr"><term>ARN viral</term>
</keywords>
<keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>SARS Virus</term>
</keywords>
<keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ARN viral</term>
<term>Virus du SRAS</term>
</keywords>
<keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>SARS Virus</term>
</keywords>
<keywords scheme="MESH" qualifier="isolement et purification" xml:lang="fr"><term>Virus du SRAS</term>
</keywords>
<keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>Reverse Transcriptase Polymerase Chain Reaction</term>
</keywords>
<keywords scheme="MESH" qualifier="virologie" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term>
</keywords>
<keywords scheme="MESH" qualifier="virology" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term>
</keywords>
<keywords scheme="MESH" xml:lang="en"><term>Case-Control Studies</term>
<term>Fluorescence</term>
<term>Humans</term>
<term>Reference Values</term>
<term>Sensitivity and Specificity</term>
</keywords>
<keywords scheme="MESH" xml:lang="fr"><term>Amorces ADN</term>
<term>Fluorescence</term>
<term>Humains</term>
<term>RT-PCR</term>
<term>Sensibilité et spécificité</term>
<term>Valeurs de référence</term>
<term>Études cas-témoins</term>
</keywords>
</textClass>
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</teiHeader>
<front><div type="abstract" xml:lang="en"><p><bold>Aim:</bold>
 SARS-associated coronavirus (<italic>SARS-CoV</italic>
) has been confirmed as the pathogen for severe acute respiratory syndrome (SARS). The aim of our study was to construct a sensitive and specific real-time quantitative fluorescent (QF) reverse transcriptase (RT)-PCR method for the detection of <italic>SARS-CoV</italic>
 RNA.</p>
<p><bold>Methods:</bold>
 Stored blood specimens from 44 patients with confirmed SARS were used along with blood samples from two sets of controls, 30 healthy volunteers who had no contact with SARS patients, and 30 healthy doctors and nurses who had contact with SARS patients but were without symptoms of SARS. Two pairs of primers were synthesized by the Shanghai Sangon Company according to SARS-CoV BJ01 strain sequence (AY278488), and then a pair of primers were designed and compared with a pair of primers published by WHO.</p>
<p><bold>Results:</bold>
 Using serial dilutions of <italic>SARS-CoV</italic>
, the 44 blood samples from SARS patients specimens were tested. Using a 0.01% dilution of <italic>SARS-CoV</italic>
, all 44 clinical samples tested positive in our assay. In comparison, using a 0.1% dilution of <italic>SARS-CoV</italic>
, 26 of the 44 samples tested positive using the WHO primers. In the QF-RT-PCR assay, there was a linear amplification from 100 copies to 10<sup>8</sup>
 copies of the control RNA per RT-PCR and at least 10 copies, and sometimes even 1 copy, of target RNA tested positive in our assay.</p>
<p><bold>Conclusion:</bold>
 The primer we developed is sufficiently sensitive and specific to diagnose symptomatic <italic>SARS-CoV</italic>
 infections and for monitoring virus load.</p>
</div>
</front>
<back><div1 type="bibliography"><listBibl><biblStruct></biblStruct>
<biblStruct></biblStruct>
<biblStruct><analytic><author><name sortKey="Peiris, Jsm" uniqKey="Peiris J">JSM Peiris</name>
</author>
<author><name sortKey="Lai, St" uniqKey="Lai S">ST Lai</name>
</author>
<author><name sortKey="Poon, Llm" uniqKey="Poon L">LLM Poon</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Ksiazek, Tg" uniqKey="Ksiazek T">TG Ksiazek</name>
</author>
<author><name sortKey="Erdman, D" uniqKey="Erdman D">D Erdman</name>
</author>
<author><name sortKey="Goldsmith, Cs" uniqKey="Goldsmith C">CS Goldsmith</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Drosten, C" uniqKey="Drosten C">C Drosten</name>
</author>
<author><name sortKey="Gunther, S" uniqKey="Gunther S">S Gunther</name>
</author>
<author><name sortKey="Preiser, W" uniqKey="Preiser W">W Preiser</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Rota, Pa" uniqKey="Rota P">PA Rota</name>
</author>
<author><name sortKey="Oberste, Ms" uniqKey="Oberste M">MS Oberste</name>
</author>
<author><name sortKey="Monroe, Ss" uniqKey="Monroe S">SS Monroe</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Marra, Ma" uniqKey="Marra M">MA Marra</name>
</author>
<author><name sortKey="Jones, Sj" uniqKey="Jones S">SJ Jones</name>
</author>
<author><name sortKey="Astell, Cr" uniqKey="Astell C">CR Astell</name>
</author>
</analytic>
</biblStruct>
<biblStruct><analytic><author><name sortKey="Qin, E" uniqKey="Qin E">E Qin</name>
</author>
<author><name sortKey="Zhu, Q" uniqKey="Zhu Q">Q Zhu</name>
</author>
<author><name sortKey="Wang, J" uniqKey="Wang J">J Wang</name>
</author>
</analytic>
</biblStruct>
<biblStruct></biblStruct>
<biblStruct></biblStruct>
<biblStruct></biblStruct>
</listBibl>
</div1>
</back>
</TEI>
<affiliations><list><country><li>République populaire de Chine</li>
</country>
<settlement><li>Pékin</li>
</settlement>
</list>
<tree><country name="République populaire de Chine"><noRegion><name sortKey="Chen, Weijun" sort="Chen, Weijun" uniqKey="Chen W" first="Weijun" last="Chen">Weijun Chen</name>
</noRegion>
<name sortKey="Chen, Weijun" sort="Chen, Weijun" uniqKey="Chen W" first="Weijun" last="Chen">Weijun Chen</name>
<name sortKey="Fang, Jianqiu" sort="Fang, Jianqiu" uniqKey="Fang J" first="Jianqiu" last="Fang">Jianqiu Fang</name>
<name sortKey="Fang, Jianqiu" sort="Fang, Jianqiu" uniqKey="Fang J" first="Jianqiu" last="Fang">Jianqiu Fang</name>
<name sortKey="Gan, Haixue" sort="Gan, Haixue" uniqKey="Gan H" first="Haixue" last="Gan">Haixue Gan</name>
<name sortKey="He, Bo" sort="He, Bo" uniqKey="He B" first="Bo" last="He">Bo He</name>
<name sortKey="Huang, Shengyong" sort="Huang, Shengyong" uniqKey="Huang S" first="Shengyong" last="Huang">Shengyong Huang</name>
<name sortKey="Lin, Lin" sort="Lin, Lin" uniqKey="Lin L" first="Lin" last="Lin">Lin Lin</name>
<name sortKey="Meng, Shufang" sort="Meng, Shufang" uniqKey="Meng S" first="Shufang" last="Meng">Shufang Meng</name>
<name sortKey="Mu, Feng" sort="Mu, Feng" uniqKey="Mu F" first="Feng" last="Mu">Feng Mu</name>
<name sortKey="Mu, Feng" sort="Mu, Feng" uniqKey="Mu F" first="Feng" last="Mu">Feng Mu</name>
<name sortKey="Mu, Jingsong" sort="Mu, Jingsong" uniqKey="Mu J" first="Jingsong" last="Mu">Jingsong Mu</name>
<name sortKey="Wang, Jian" sort="Wang, Jian" uniqKey="Wang J" first="Jian" last="Wang">Jian Wang</name>
<name sortKey="Wang, Jian" sort="Wang, Jian" uniqKey="Wang J" first="Jian" last="Wang">Jian Wang</name>
<name sortKey="Wen, Jie" sort="Wen, Jie" uniqKey="Wen J" first="Jie" last="Wen">Jie Wen</name>
<name sortKey="Wen, Jie" sort="Wen, Jie" uniqKey="Wen J" first="Jie" last="Wen">Jie Wen</name>
<name sortKey="Xu, Zuyuan" sort="Xu, Zuyuan" uniqKey="Xu Z" first="Zuyuan" last="Xu">Zuyuan Xu</name>
<name sortKey="Xu, Zuyuan" sort="Xu, Zuyuan" uniqKey="Xu Z" first="Zuyuan" last="Xu">Zuyuan Xu</name>
<name sortKey="Yang, Ling" sort="Yang, Ling" uniqKey="Yang L" first="Ling" last="Yang">Ling Yang</name>
<name sortKey="Yang, Ling" sort="Yang, Ling" uniqKey="Yang L" first="Ling" last="Yang">Ling Yang</name>
</country>
</tree>
</affiliations>
</record>

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Real-Time Quantitative Fluorescent Reverse Transcriptase-PCR for Detection of Severe Acute Respiratory Syndrome-Associated Coronavirus RNA

Real-Time Quantitative Fluorescent Reverse Transcriptase-PCR for Detection of Severe Acute Respiratory Syndrome-Associated Coronavirus RNA

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