Narcissus yellow stripe virus and Narcissus mosaic virus detection in Narcissus via multiplex TaqMan-based reverse transcription-PCR assay
| Autor: | F.R. Liao, J.F. Ma, Zujian Wu, Jing Jin, J.G. Shen, G.H. Xie, X.H. Chen, Fangluan Gao, W. Cai |
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| Rok vydání: | 2017 |
| Předmět: |
0301 basic medicine
food.ingredient Narcissus mosaic virus Potyvirus Narcissus yellow stripe virus Sensitivity and Specificity Applied Microbiology and Biotechnology law.invention 03 medical and health sciences food law Plant virus TaqMan Multiplex Polymerase chain reaction DNA Primers biology Reverse Transcriptase Polymerase Chain Reaction Reproducibility of Results Narcissus Reverse Transcription General Medicine biology.organism_classification Virology Molecular biology Reverse transcription polymerase chain reaction 030104 developmental biology Multiplex Polymerase Chain Reaction Biotechnology |
| Zdroj: | Journal of Applied Microbiology. 122:1299-1309 |
| ISSN: | 1364-5072 |
| Popis: | Aims Development of a multiplex TaqMan RT-qPCR assay to simultaneously detect Narcissus yellow stripe virus (NYSV) and Narcissus mosaic virus (NMV), frequently causing mixed narcissus infection. Feasibility verification was confirmed in natural samples. Methods and Results Primers and probes were designed based on the conserved CP gene regions of NYSV or NMV and their suitability for singleplex and multiplex TaqMan RT-qPCR assays as well as for conventional RT-PCR. Conventional RT-PCR, singleplex and multiplex TaqMan RT-qPCR assays proved to be NYSV and NMV specific. P-values and coefficients of variation of TaqMan RT-qPCR assays indicated high reproducibility. Significantly increased sensitivity was achieved compared to conventional RT-PCR. The detection limit of both viruses was 103 copies with superior correlation coefficients and linear standard curve responses between plasmid concentrations and Ct values. NYSV and NMV infection of narcissus leaves, petals and bulbs could successfully be detected via our multiplex RT-qPCR method at 1·25 mg. Conclusion Our multiplex TaqMan RT-qPCR assay provides rapid, specific, sensitive and reliable testing to simultaneously detect NYSV and NMV, supplying useful routine monitoring for different narcissus samples. Significance and Impact of the Study Efficient identification and discrimination of the narcissus viruses provides reliable information for scientists and conventional growers. Furthermore, it enriches the information of NYSV, NMV and other narcissus viruses. |
| Databáze: | OpenAIRE |
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