One-tube nested RT-PCR enabled by using a plastic film and its application for the rapid detection of SARS-virus.
Identifieur interne : 002E76 ( PubMed/Corpus ); précédent : 002E75; suivant : 002E77One-tube nested RT-PCR enabled by using a plastic film and its application for the rapid detection of SARS-virus.
Auteurs : Sheng-Ce Tao ; Di Jiang ; Hong-Li Lu ; Wan-Li Xing ; Yu-Xiang Zhou ; Jing ChengSource :
- Biotechnology letters [ 0141-5492 ] ; 2004.
English descriptors
- KwdEn :
- Biocompatible Materials, Equipment Design, Equipment Failure Analysis, Polyethylenes, RNA, Viral (analysis), RNA, Viral (genetics), Reproducibility of Results, Reverse Transcriptase Polymerase Chain Reaction (instrumentation), Reverse Transcriptase Polymerase Chain Reaction (methods), SARS Virus (genetics), SARS Virus (isolation & purification), Sensitivity and Specificity.
- MESH :
- chemical , analysis : RNA, Viral.
- chemical , genetics : RNA, Viral.
- chemical : Biocompatible Materials, Polyethylenes.
- genetics : SARS Virus.
- instrumentation : Reverse Transcriptase Polymerase Chain Reaction.
- isolation & purification : SARS Virus.
- methods : Reverse Transcriptase Polymerase Chain Reaction.
- Equipment Design, Equipment Failure Analysis, Reproducibility of Results, Sensitivity and Specificity.
Abstract
A general strategy based on the use of a plastic film that allows reverse transcription and nested-PCR in a single closed-tube has been developed. The reaction mixture for the second PCR amplification is quarantined in the cap of the reaction tube during the first round amplification by a piece of plastic film, and later introduced into the PCR amplicons from the first round reaction by centrifugation without opening the reaction tube. The main advantages of our method are its high sensitivity, specificity, simplicity, cost effectiveness, low risk of contamination and the ease in establishment of conditions for nested-PCR. The method has been successfully applied to the detection of the genomic RNA from SARS-CoV, the detection limit of this method is comparable to that of the conventional two-tube nested PCR system. This method could be easily adapted to the detection of other targets.
DOI: 10.1023/b:bile.0000013708.65032.f1
PubMed: 15049359
Links to Exploration step
pubmed:15049359Le document en format XML
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<author><name sortKey="Tao, Sheng Ce" sort="Tao, Sheng Ce" uniqKey="Tao S" first="Sheng-Ce" last="Tao">Sheng-Ce Tao</name>
<affiliation><nlm:affiliation>Department of Biological Sciences and Biotechnology, Tsinghua University, Haidian District, Beijing 100084, PR China.</nlm:affiliation>
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<author><name sortKey="Jiang, Di" sort="Jiang, Di" uniqKey="Jiang D" first="Di" last="Jiang">Di Jiang</name>
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<author><name sortKey="Lu, Hong Li" sort="Lu, Hong Li" uniqKey="Lu H" first="Hong-Li" last="Lu">Hong-Li Lu</name>
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<author><name sortKey="Xing, Wan Li" sort="Xing, Wan Li" uniqKey="Xing W" first="Wan-Li" last="Xing">Wan-Li Xing</name>
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<author><name sortKey="Zhou, Yu Xiang" sort="Zhou, Yu Xiang" uniqKey="Zhou Y" first="Yu-Xiang" last="Zhou">Yu-Xiang Zhou</name>
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<author><name sortKey="Cheng, Jing" sort="Cheng, Jing" uniqKey="Cheng J" first="Jing" last="Cheng">Jing Cheng</name>
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<author><name sortKey="Tao, Sheng Ce" sort="Tao, Sheng Ce" uniqKey="Tao S" first="Sheng-Ce" last="Tao">Sheng-Ce Tao</name>
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<author><name sortKey="Xing, Wan Li" sort="Xing, Wan Li" uniqKey="Xing W" first="Wan-Li" last="Xing">Wan-Li Xing</name>
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<author><name sortKey="Zhou, Yu Xiang" sort="Zhou, Yu Xiang" uniqKey="Zhou Y" first="Yu-Xiang" last="Zhou">Yu-Xiang Zhou</name>
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<author><name sortKey="Cheng, Jing" sort="Cheng, Jing" uniqKey="Cheng J" first="Jing" last="Cheng">Jing Cheng</name>
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<series><title level="j">Biotechnology letters</title>
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<profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Biocompatible Materials</term>
<term>Equipment Design</term>
<term>Equipment Failure Analysis</term>
<term>Polyethylenes</term>
<term>RNA, Viral (analysis)</term>
<term>RNA, Viral (genetics)</term>
<term>Reproducibility of Results</term>
<term>Reverse Transcriptase Polymerase Chain Reaction (instrumentation)</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>
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<front><div type="abstract" xml:lang="en">A general strategy based on the use of a plastic film that allows reverse transcription and nested-PCR in a single closed-tube has been developed. The reaction mixture for the second PCR amplification is quarantined in the cap of the reaction tube during the first round amplification by a piece of plastic film, and later introduced into the PCR amplicons from the first round reaction by centrifugation without opening the reaction tube. The main advantages of our method are its high sensitivity, specificity, simplicity, cost effectiveness, low risk of contamination and the ease in establishment of conditions for nested-PCR. The method has been successfully applied to the detection of the genomic RNA from SARS-CoV, the detection limit of this method is comparable to that of the conventional two-tube nested PCR system. This method could be easily adapted to the detection of other targets.</div>
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<Abstract><AbstractText>A general strategy based on the use of a plastic film that allows reverse transcription and nested-PCR in a single closed-tube has been developed. The reaction mixture for the second PCR amplification is quarantined in the cap of the reaction tube during the first round amplification by a piece of plastic film, and later introduced into the PCR amplicons from the first round reaction by centrifugation without opening the reaction tube. The main advantages of our method are its high sensitivity, specificity, simplicity, cost effectiveness, low risk of contamination and the ease in establishment of conditions for nested-PCR. The method has been successfully applied to the detection of the genomic RNA from SARS-CoV, the detection limit of this method is comparable to that of the conventional two-tube nested PCR system. This method could be easily adapted to the detection of other targets.</AbstractText>
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