Preparation of His-tagged armored RNA phage particles as a control for real-time reverse transcription-PCR detection of severe acute respiratory syndrome coronavirus.
Identifieur interne : 003F49 ( Main/Merge ); précédent : 003F48; suivant : 003F50Preparation of His-tagged armored RNA phage particles as a control for real-time reverse transcription-PCR detection of severe acute respiratory syndrome coronavirus.
Auteurs : Yangjian Cheng [République populaire de Chine] ; Jianjun Niu ; Yongyou Zhang ; Jianwei Huang ; Qingge LiSource :
- Journal of clinical microbiology [ 0095-1137 ] ; 2006.
Descripteurs français
- KwdFr :
- MESH :
- isolement et purification : Virus du SRAS.
- ARN viral, Histidine, Phages à ARN, RT-PCR, Sensibilité et spécificité.
English descriptors
- KwdEn :
- MESH :
- chemical , chemistry : Histidine.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction.
- RNA Phages, RNA, Viral, Sensitivity and Specificity.
Abstract
Armored RNA has been increasingly used as both an external and internal positive control in nucleic acid-based assays for RNA virus. In order to facilitate armored RNA purification, a His6 tag was introduced into the loop region of the MS2 coat protein, which allows the exposure of multiple His tags on the surface during armored RNA assembly. The His-tagged armored RNA particles were purified to homogeneity and verified to be free of DNA contamination in a single run of affinity chromatography. A fragment of severe acute respiratory syndrome coronavirus (SARS-CoV) genome targeted for SARS-CoV detection was chosen for an external positive control preparation. A plant-specific gene sequence was chosen for a universal noncompetitive internal positive control preparation. Both controls were purified by Co2+ affinity chromatography and were included in a real-time reverse transcription-PCR assay for SARS-CoV. The noncompetitive internal positive control can be added to clinical samples before RNA extraction and enables the identification of potential inhibitive effects without interfering with target amplification. The external control could be used for the quantification of viral loads in clinical samples.
DOI: 10.1128/JCM.00713-06
PubMed: 17021082
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pubmed:17021082Le document en format XML
<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Preparation of His-tagged armored RNA phage particles as a control for real-time reverse transcription-PCR detection of severe acute respiratory syndrome coronavirus.</title><author><name sortKey="Cheng, Yangjian" sort="Cheng, Yangjian" uniqKey="Cheng Y" first="Yangjian" last="Cheng">Yangjian Cheng</name><affiliation wicri:level="1"><nlm:affiliation>Department of Biomedical Sciences, Xiamen University, South Road of Siming 422, Xiamen 361005, China.</nlm:affiliation><country xml:lang="fr">République populaire de Chine</country><wicri:regionArea>Department of Biomedical Sciences, Xiamen University, South Road of Siming 422, Xiamen 361005</wicri:regionArea><wicri:noRegion>Xiamen 361005</wicri:noRegion></affiliation></author><author><name sortKey="Niu, Jianjun" sort="Niu, Jianjun" uniqKey="Niu J" first="Jianjun" last="Niu">Jianjun Niu</name></author><author><name sortKey="Zhang, Yongyou" sort="Zhang, Yongyou" uniqKey="Zhang Y" first="Yongyou" last="Zhang">Yongyou Zhang</name></author><author><name sortKey="Huang, Jianwei" sort="Huang, Jianwei" uniqKey="Huang J" first="Jianwei" last="Huang">Jianwei Huang</name></author><author><name sortKey="Li, Qingge" sort="Li, Qingge" uniqKey="Li Q" first="Qingge" last="Li">Qingge Li</name></author></titleStmt><publicationStmt><idno type="wicri:source">PubMed</idno><date when="2006">2006</date><idno type="RBID">pubmed:17021082</idno><idno type="pmid">17021082</idno><idno type="doi">10.1128/JCM.00713-06</idno><idno type="wicri:Area/PubMed/Corpus">002071</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Corpus" wicri:corpus="PubMed">002071</idno><idno type="wicri:Area/PubMed/Curation">002071</idno><idno type="wicri:explorRef" wicri:stream="PubMed" wicri:step="Curation">002071</idno><idno type="wicri:Area/PubMed/Checkpoint">002061</idno><idno type="wicri:explorRef" wicri:stream="Checkpoint" wicri:step="PubMed">002061</idno><idno 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361005</wicri:regionArea><wicri:noRegion>Xiamen 361005</wicri:noRegion></affiliation></author><author><name sortKey="Niu, Jianjun" sort="Niu, Jianjun" uniqKey="Niu J" first="Jianjun" last="Niu">Jianjun Niu</name></author><author><name sortKey="Zhang, Yongyou" sort="Zhang, Yongyou" uniqKey="Zhang Y" first="Yongyou" last="Zhang">Yongyou Zhang</name></author><author><name sortKey="Huang, Jianwei" sort="Huang, Jianwei" uniqKey="Huang J" first="Jianwei" last="Huang">Jianwei Huang</name></author><author><name sortKey="Li, Qingge" sort="Li, Qingge" uniqKey="Li Q" first="Qingge" last="Li">Qingge Li</name></author></analytic><series><title level="j">Journal of clinical microbiology</title><idno type="ISSN">0095-1137</idno><imprint><date when="2006" type="published">2006</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Histidine (chemistry)</term><term>RNA Phages</term><term>RNA, Viral</term><term>Reverse Transcriptase Polymerase Chain Reaction (methods)</term><term>SARS Virus (isolation & purification)</term><term>Sensitivity and Specificity</term></keywords><keywords scheme="KwdFr" xml:lang="fr"><term>ARN viral</term><term>Histidine ()</term><term>Phages à ARN</term><term>RT-PCR ()</term><term>Sensibilité et spécificité</term><term>Virus du SRAS (isolement et purification)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="chemistry" xml:lang="en"><term>Histidine</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" xml:lang="en"><term>RNA Phages</term><term>RNA, Viral</term><term>Sensitivity and Specificity</term></keywords><keywords 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In order to facilitate armored RNA purification, a His6 tag was introduced into the loop region of the MS2 coat protein, which allows the exposure of multiple His tags on the surface during armored RNA assembly. The His-tagged armored RNA particles were purified to homogeneity and verified to be free of DNA contamination in a single run of affinity chromatography. A fragment of severe acute respiratory syndrome coronavirus (SARS-CoV) genome targeted for SARS-CoV detection was chosen for an external positive control preparation. A plant-specific gene sequence was chosen for a universal noncompetitive internal positive control preparation. Both controls were purified by Co2+ affinity chromatography and were included in a real-time reverse transcription-PCR assay for SARS-CoV. The noncompetitive internal positive control can be added to clinical samples before RNA extraction and enables the identification of potential inhibitive effects without interfering with target amplification. The external control could be used for the quantification of viral loads in clinical samples.</div></front></TEI></record>Pour manipuler ce document sous Unix (Dilib)
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