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Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR
Seng, EK; Fang, Q; Lam, TJ; Sin, YM; Seng, EK, Natl Univ Singapore, Dept Biol Sci, Block S1A,05-02 Virol Lab,14 Sci Dr 4, Singapore 117543, Singapore
2004-06-15
Source PublicationJOURNAL OF VIROLOGICAL METHODS
ISSN0166-0934
Volume118Issue:2Pages:111-122
AbstractA rapid, sensitive and highly specific detection method for Aquareovirus based on reverse-transcription polymerase chain reaction (RT-PCR) was developed. Based on multiple sequence alignment of the cloned sequences of a local isolates, the Threadfin reovirus (TFV) and Guppy reovirus (GPV) with Grass carp reovirus (GCRV), a pair of degenerate primers was selected carefully and synthesized. Using this primer combination, only one specific product, approximately 450 bp in length was obtained when RT-PCR was carried out using the genomic double-stranded RNA (dsRNA) of TFV, GPV and GCRV. Similar results were also obtained when Chum salmon reovirus (CSRV) and Striped bass reovirus (SBRV) dsRNA were used as templates. No products were observed when nucleic acids other than the dsRNA of the aquareoviruses described above were used as RT-PCR templates. This technique could detect not only TFV but also GPV and GCRV in low titer virus-infected cell cultured cells. Furthermore, this method has also been shown to be able to diagnose GPV-infected guppy (Poecilia reticulata) that exhibit clinical symptoms as well as GPV-carrier guppy. Collectively, these results showed that the RT-PCR amplification method using specific degenerate primers described below is very useful for rapid and accurate detection of a variety of aquareovirus strains isolated from different host species and origin. (C) 2004 Elsevier B.V. All rights reserved.; A rapid, sensitive and highly specific detection method for Aquareovirus based on reverse-transcription polymerase chain reaction (RT-PCR) was developed. Based on multiple sequence alignment of the cloned sequences of a local isolates, the Threadfin reovirus (TFV) and Guppy reovirus (GPV) with Grass carp reovirus (GCRV), a pair of degenerate primers was selected carefully and synthesized. Using this primer combination, only one specific product, approximately 450 bp in length was obtained when RT-PCR was carried out using the genomic double-stranded RNA (dsRNA) of TFV, GPV and GCRV. Similar results were also obtained when Chum salmon reovirus (CSRV) and Striped bass reovirus (SBRV) dsRNA were used as templates. No products were observed when nucleic acids other than the dsRNA of the aquareoviruses described above were used as RT-PCR templates. This technique could detect not only TFV but also GPV and GCRV in low titer virus-infected cell cultured cells. Furthermore, this method has also been shown to be able to diagnose GPV-infected guppy (Poecilia reticulata) that exhibit clinical symptoms as well as GPV-carrier guppy. Collectively, these results showed that the RT-PCR amplification method using specific degenerate primers described below is very useful for rapid and accurate detection of a variety of aquareovirus strains isolated from different host species and origin. (C) 2004 Elsevier B.V. All rights reserved.
SubtypeArticle
KeywordAquareovirus Rt-pcr Degenerate Primers Threadfin Reovirus
DepartmentNatl Univ Singapore, Dept Biol Sci, Singapore 117543, Singapore; CAS, Wuhan Inst Virol, Inst Hydrobiol, Wuhan 430071, Hubei, Peoples R China; Teo Way Yong & Sons Pte Ltd, Singapore Fish Breeding & Immunizat Ctr, Singapore, Singapore
Subject AreaBiochemical Research Methods ; Biotechnology & Applied Microbiology ; Virology
DOI10.1016/j.jviromet.2004.01.023
WOS HeadingsScience & Technology ; Life Sciences & Biomedicine
Indexed BySCI
Language英语
WOS Research AreaBiochemistry & Molecular Biology ; Biotechnology & Applied Microbiology ; Virology
WOS SubjectBiochemical Research Methods ; Biotechnology & Applied Microbiology ; Virology
WOS IDWOS:000221207300005
WOS KeywordRNA BLOT HYBRIDIZATION ; CELL-CULTURE ; IDENTIFICATION ; VIRUSES ; GENOGROUP ; ROTAVIRUS ; INFECTION ; REOVIRUS ; PROBE
Citation statistics
Cited Times:21[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Identifierhttp://ir.ihb.ac.cn/handle/152342/9496
Collection期刊论文
Corresponding AuthorSeng, EK, Natl Univ Singapore, Dept Biol Sci, Block S1A,05-02 Virol Lab,14 Sci Dr 4, Singapore 117543, Singapore
Affiliation1.Natl Univ Singapore, Dept Biol Sci, Singapore 117543, Singapore
2.CAS, Wuhan Inst Virol, Inst Hydrobiol, Wuhan 430071, Hubei, Peoples R China
3.Teo Way Yong & Sons Pte Ltd, Singapore Fish Breeding & Immunizat Ctr, Singapore, Singapore
Recommended Citation
GB/T 7714
Seng, EK,Fang, Q,Lam, TJ,et al. Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR[J]. JOURNAL OF VIROLOGICAL METHODS,2004,118(2):111-122.
APA Seng, EK,Fang, Q,Lam, TJ,Sin, YM,&Seng, EK, Natl Univ Singapore, Dept Biol Sci, Block S1A,05-02 Virol Lab,14 Sci Dr 4, Singapore 117543, Singapore.(2004).Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR.JOURNAL OF VIROLOGICAL METHODS,118(2),111-122.
MLA Seng, EK,et al."Development of a rapid, sensitive and specific diagnostic assay for fish Aquareovirus based on RT-PCR".JOURNAL OF VIROLOGICAL METHODS 118.2(2004):111-122.
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