Development of Taqman RT-nested PCR system for clinical SARS-CoV detection.
Identifieur interne : 005388 ( Main/Exploration ); précédent : 005387; suivant : 005389Development of Taqman RT-nested PCR system for clinical SARS-CoV detection.
Auteurs : Qingfa Wu [République populaire de Chine] ; Zuyuan Xu ; Tian Wei ; Haipang Zeng ; Jingxiang Li ; Haixue Gang ; Min Sun ; Fangbo Jiang ; Xiang Wang ; Wei Dong ; Ling Yang ; Jian WangSource :
- Journal of virological methods [ 0166-0934 ] ; 2004.
Descripteurs français
- KwdFr :
- ADN viral (génétique), Adolescent, Adulte, Adulte d'âge moyen, Amorces ADN (génétique), Chine, Enfant, Femelle, Génome viral, Humains, Mâle, RT-PCR (), Sensibilité et spécificité, Sujet âgé, Syndrome respiratoire aigu sévère (diagnostic), Syndrome respiratoire aigu sévère (virologie), Séquence nucléotidique, Virologie (), Virus du SRAS (génétique), Virus du SRAS (isolement et purification).
- MESH :
- diagnostic : Syndrome respiratoire aigu sévère.
- génétique : ADN viral, Amorces ADN, Virus du SRAS.
- isolement et purification : Virus du SRAS.
- virologie : Syndrome respiratoire aigu sévère.
- Adolescent, Adulte, Adulte d'âge moyen, Chine, Enfant, Femelle, Génome viral, Humains, Mâle, RT-PCR, Sensibilité et spécificité, Sujet âgé, Séquence nucléotidique, Virologie.
- Wicri :
- geographic : République populaire de Chine.
English descriptors
- KwdEn :
- Adolescent, Adult, Aged, Base Sequence, Child, China, DNA Primers (genetics), DNA, Viral (genetics), Female, Genome, Viral, Humans, Male, Middle Aged, Reverse Transcriptase Polymerase Chain Reaction (methods), Reverse Transcriptase Polymerase Chain Reaction (statistics & numerical data), SARS Virus (genetics), SARS Virus (isolation & purification), Sensitivity and Specificity, Severe Acute Respiratory Syndrome (diagnosis), Severe Acute Respiratory Syndrome (virology), Virology (methods), Virology (statistics & numerical data).
- MESH :
- chemical , genetics : DNA Primers, DNA, Viral.
- geographic : China.
- diagnosis : Severe Acute Respiratory Syndrome.
- genetics : SARS Virus.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction, Virology.
- statistics & numerical data : Reverse Transcriptase Polymerase Chain Reaction, Virology.
- virology : Severe Acute Respiratory Syndrome.
- Adolescent, Adult, Aged, Base Sequence, Child, Female, Genome, Viral, Humans, Male, Middle Aged, Sensitivity and Specificity.
Abstract
Severe acute respiratory syndrome (SARS) is an acute newly emerged infectious respiratory illness. The etiologic agent of SARS was named 'SARS-associated coronavirus' (SARS-CoV) that can be detected with reverse transcription-polymerase chain reaction (RT-PCR) assays. In this study, 12 sets of nested primers covering the SARS-CoV genome have been screened and showed sufficient sensitivity to detect SARS-CoV in RNA isolated from virus cultured in Vero 6 cells. To optimize further the reaction condition of those nested primers sets, seven sets of nested primers have been chosen to compare their reverse transcribed efficiency with specific and random primers, which is useful to combine RT with the first round of PCR into a one-step RT-PCR. Based on the sensitivity and simplicity of results, the no. 73 primer set was chosen as the candidate primer set for clinical diagnoses. To specify the amplicon to minimize false positive results, a Taqman RT-nested PCR system of no. 73 nested primer set was developed. Through investigations on a test panel of whole blood obtained from 30 SARS patients and 9 control persons, the specificity and sensitivity of the Taqman RT-nested PCR system was found to be 100 and 83%, respectively, which suggests that the method is a promising one to diagnose SARS in early stages.
DOI: 10.1016/j.jviromet.2004.02.011
PubMed: 15109816
Affiliations:
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sort="Zeng, Haipang" uniqKey="Zeng H" first="Haipang" last="Zeng">Haipang Zeng</name></author><author><name sortKey="Li, Jingxiang" sort="Li, Jingxiang" uniqKey="Li J" first="Jingxiang" last="Li">Jingxiang Li</name></author><author><name sortKey="Gang, Haixue" sort="Gang, Haixue" uniqKey="Gang H" first="Haixue" last="Gang">Haixue Gang</name></author><author><name sortKey="Sun, Min" sort="Sun, Min" uniqKey="Sun M" first="Min" last="Sun">Min Sun</name></author><author><name sortKey="Jiang, Fangbo" sort="Jiang, Fangbo" uniqKey="Jiang F" first="Fangbo" last="Jiang">Fangbo Jiang</name></author><author><name sortKey="Wang, Xiang" sort="Wang, Xiang" uniqKey="Wang X" first="Xiang" last="Wang">Xiang Wang</name></author><author><name sortKey="Dong, Wei" sort="Dong, Wei" uniqKey="Dong W" first="Wei" last="Dong">Wei Dong</name></author><author><name sortKey="Yang, Ling" sort="Yang, Ling" uniqKey="Yang L" first="Ling" last="Yang">Ling Yang</name></author><author><name sortKey="Wang, Jian" 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Xu</name></author><author><name sortKey="Wei, Tian" sort="Wei, Tian" uniqKey="Wei T" first="Tian" last="Wei">Tian Wei</name></author><author><name sortKey="Zeng, Haipang" sort="Zeng, Haipang" uniqKey="Zeng H" first="Haipang" last="Zeng">Haipang Zeng</name></author><author><name sortKey="Li, Jingxiang" sort="Li, Jingxiang" uniqKey="Li J" first="Jingxiang" last="Li">Jingxiang Li</name></author><author><name sortKey="Gang, Haixue" sort="Gang, Haixue" uniqKey="Gang H" first="Haixue" last="Gang">Haixue Gang</name></author><author><name sortKey="Sun, Min" sort="Sun, Min" uniqKey="Sun M" first="Min" last="Sun">Min Sun</name></author><author><name sortKey="Jiang, Fangbo" sort="Jiang, Fangbo" uniqKey="Jiang F" first="Fangbo" last="Jiang">Fangbo Jiang</name></author><author><name sortKey="Wang, Xiang" sort="Wang, Xiang" uniqKey="Wang X" first="Xiang" last="Wang">Xiang Wang</name></author><author><name sortKey="Dong, Wei" sort="Dong, Wei" uniqKey="Dong W" first="Wei" last="Dong">Wei Dong</name></author><author><name sortKey="Yang, Ling" sort="Yang, Ling" uniqKey="Yang L" first="Ling" last="Yang">Ling Yang</name></author><author><name sortKey="Wang, Jian" sort="Wang, Jian" uniqKey="Wang J" first="Jian" last="Wang">Jian Wang</name></author></analytic><series><title level="j">Journal of virological methods</title><idno type="ISSN">0166-0934</idno><imprint><date when="2004" type="published">2004</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Adolescent</term><term>Adult</term><term>Aged</term><term>Base Sequence</term><term>Child</term><term>China</term><term>DNA Primers (genetics)</term><term>DNA, Viral (genetics)</term><term>Female</term><term>Genome, Viral</term><term>Humans</term><term>Male</term><term>Middle Aged</term><term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term><term>Reverse Transcriptase Polymerase Chain Reaction (statistics & numerical data)</term><term>SARS Virus (genetics)</term><term>SARS Virus (isolation & purification)</term><term>Sensitivity and Specificity</term><term>Severe Acute Respiratory Syndrome (diagnosis)</term><term>Severe Acute Respiratory Syndrome (virology)</term><term>Virology (methods)</term><term>Virology (statistics & numerical data)</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ADN viral (génétique)</term><term>Adolescent</term><term>Adulte</term><term>Adulte d'âge moyen</term><term>Amorces ADN (génétique)</term><term>Chine</term><term>Enfant</term><term>Femelle</term><term>Génome viral</term><term>Humains</term><term>Mâle</term><term>RT-PCR ()</term><term>Sensibilité et spécificité</term><term>Sujet âgé</term><term>Syndrome respiratoire aigu sévère (diagnostic)</term><term>Syndrome respiratoire aigu sévère (virologie)</term><term>Séquence nucléotidique</term><term>Virologie ()</term><term>Virus du SRAS (génétique)</term><term>Virus du SRAS (isolement et purification)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA Primers</term><term>DNA, Viral</term></keywords><keywords scheme="MESH" type="geographic" xml:lang="en"><term>China</term></keywords><keywords scheme="MESH" qualifier="diagnosis" xml:lang="en"><term>Severe Acute Respiratory Syndrome</term></keywords><keywords scheme="MESH" qualifier="diagnostic" xml:lang="fr"><term>Syndrome respiratoire aigu sévère</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>SARS Virus</term></keywords><keywords scheme="MESH" qualifier="génétique" xml:lang="fr"><term>ADN viral</term><term>Amorces ADN</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><term>Virology</term></keywords><keywords scheme="MESH" qualifier="statistics & numerical data" xml:lang="en"><term>Reverse Transcriptase Polymerase Chain Reaction</term><term>Virology</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>Adolescent</term><term>Adult</term><term>Aged</term><term>Base Sequence</term><term>Child</term><term>Female</term><term>Genome, Viral</term><term>Humans</term><term>Male</term><term>Middle Aged</term><term>Sensitivity and Specificity</term></keywords><keywords scheme="MESH" xml:lang="fr"><term>Adolescent</term><term>Adulte</term><term>Adulte d'âge moyen</term><term>Chine</term><term>Enfant</term><term>Femelle</term><term>Génome viral</term><term>Humains</term><term>Mâle</term><term>RT-PCR</term><term>Sensibilité et spécificité</term><term>Sujet âgé</term><term>Séquence nucléotidique</term><term>Virologie</term></keywords><keywords scheme="Wicri" type="geographic" xml:lang="fr"><term>République populaire de Chine</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Severe acute respiratory syndrome (SARS) is an acute newly emerged infectious respiratory illness. The etiologic agent of SARS was named 'SARS-associated coronavirus' (SARS-CoV) that can be detected with reverse transcription-polymerase chain reaction (RT-PCR) assays. In this study, 12 sets of nested primers covering the SARS-CoV genome have been screened and showed sufficient sensitivity to detect SARS-CoV in RNA isolated from virus cultured in Vero 6 cells. To optimize further the reaction condition of those nested primers sets, seven sets of nested primers have been chosen to compare their reverse transcribed efficiency with specific and random primers, which is useful to combine RT with the first round of PCR into a one-step RT-PCR. Based on the sensitivity and simplicity of results, the no. 73 primer set was chosen as the candidate primer set for clinical diagnoses. To specify the amplicon to minimize false positive results, a Taqman RT-nested PCR system of no. 73 nested primer set was developed. Through investigations on a test panel of whole blood obtained from 30 SARS patients and 9 control persons, the specificity and sensitivity of the Taqman RT-nested PCR system was found to be 100 and 83%, respectively, which suggests that the method is a promising one to diagnose SARS in early stages.</div></front></TEI><affiliations><list><country><li>République populaire de Chine</li></country><settlement><li>Pékin</li></settlement></list><tree><noCountry><name sortKey="Dong, Wei" sort="Dong, Wei" uniqKey="Dong W" first="Wei" last="Dong">Wei Dong</name><name sortKey="Gang, Haixue" sort="Gang, Haixue" uniqKey="Gang H" first="Haixue" last="Gang">Haixue Gang</name><name sortKey="Jiang, Fangbo" sort="Jiang, Fangbo" uniqKey="Jiang F" first="Fangbo" last="Jiang">Fangbo Jiang</name><name sortKey="Li, Jingxiang" sort="Li, Jingxiang" uniqKey="Li J" first="Jingxiang" last="Li">Jingxiang Li</name><name sortKey="Sun, Min" sort="Sun, Min" uniqKey="Sun M" first="Min" last="Sun">Min Sun</name><name sortKey="Wang, Jian" sort="Wang, Jian" uniqKey="Wang J" first="Jian" last="Wang">Jian Wang</name><name sortKey="Wang, Xiang" sort="Wang, Xiang" uniqKey="Wang X" first="Xiang" last="Wang">Xiang Wang</name><name sortKey="Wei, Tian" sort="Wei, Tian" uniqKey="Wei T" first="Tian" last="Wei">Tian Wei</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="Zeng, Haipang" sort="Zeng, Haipang" uniqKey="Zeng H" first="Haipang" last="Zeng">Haipang Zeng</name></noCountry><country name="République populaire de Chine"><noRegion><name sortKey="Wu, Qingfa" sort="Wu, Qingfa" uniqKey="Wu Q" first="Qingfa" last="Wu">Qingfa Wu</name></noRegion></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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