A One-Step Real-Time RT-PCR Assay for the Detection and Quantitation of Sugarcane Streak Mosaic Virus.
Sugarcane mosaic disease is caused by the Sugarcane streak mosaic virus (SCSMV; genus Poacevirus, family Potyviridae) which is common in some Asian countries. Here, we established a protocol of a one-step real-time quantitative reverse transcription PCR (real-time qRT-PCR) using the TaqMan probe for...
| Publicado en: | BioMed Research International Vol. 2015; pp. 1 - 10 |
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| Autores principales: | , , , , , |
| Formato: | pictorial research tables/charts Journal Article |
| Publicado: |
Wiley-Blackwell
6/22/2015
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| Acceso en línea: | Ver este registro en EBSCOhost |
| fields | @attributes: recordID: 1 pdfLink: plink: https://search.ebscohost.com/login.aspx?direct=true&db=ccm&AN=109274653&site=ehost-live header: @attributes: shortDbName: ccm uiTerm: 109274653 longDbName: CINAHL Complete uiTag: AN controlInfo: bkinfo: dissinfo: jinfo: jid: 23146133 FT2T jtl: BioMed Research International issn: 23146133 maglogo: N pubinfo: dt: 6/22/2015 vid: 2015 pid: 480 pub: Wiley-Blackwell place: Malden, Massachusetts artinfo: ui: 109274653 109274653 109274653 10.1155/2015/569131 109274653 ppf: 1 ppct: 9 formats: fmt: @attributes: type: P tig: atl: A One-Step Real-Time RT-PCR Assay for the Detection and Quantitation of Sugarcane Streak Mosaic Virus. aug: au: Fu, Wei-Lin Sun, Sheng-Ren Fu, Hua-Ying Chen, Ru-Kai Su, Jin-Wei Gao, San-Ji affil: Key Laboratory of Sugarcane Biology and Genetic Breeding, Ministry of Agriculture, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China sug: subj: Plant Diseases Genetics Virus Diseases Polymerase Chain Reaction Methods Gene Expression Evaluation RNA Probes Analytic Sample Preparation Methods In Vitro Studies Sequence Analysis Microbial Culture and Sensitivity Tests Descriptive Statistics Data Analysis Software Fluorescent Dyes Funding Source Evolution Models, Theoretical ab: Sugarcane mosaic disease is caused by the Sugarcane streak mosaic virus (SCSMV; genus Poacevirus, family Potyviridae) which is common in some Asian countries. Here, we established a protocol of a one-step real-time quantitative reverse transcription PCR (real-time qRT-PCR) using the TaqMan probe for the detection of SCSMV in sugarcane. Primers and probes were designed within the conserved region of the SCSMV coat protein (CP) gene sequences. Standard single-stranded RNA (ssRNA) generated by PCR-based gene transcripts of recombinant pGEM-CP plasmid in vitro and total RNA extracted from SCSMV-infected sugarcane were used as templates of qRT-PCR. We further performed a sensitivity assay to show that the detection limit of the assay was 100 copies of ssRNA and 2 pg of total RNA with good reproducibility. The values obtained were approximately 100-fold more sensitive than those of the conventional RT-PCR. A higher incidence (68.6%) of SCSMV infection was detected by qRT-PCR than that (48.6%) with conventional RT-PCR in samples showing mosaic symptoms. SCSMV-free samples were verified by infection with Sugarcane mosaic virus (SCMV) or Sorghum mosaic virus (SrMV) or a combination of both. The developed qRT-PCR assay may become an alternative molecular tool for an economical, rapid, and efficient detection and quantification of SCSMV. pubtype: Academic Journal doctype: pictorial research tables/charts Journal Article ougenre: Article language: English refInfo: holdings: @attributes: islocal: N |
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