Articles
A one step real-time RT-PCR assay for the quantitation of carnation mottle virus (CarMV)
Article number
1467_28
Pages
233 – 240
Language
English
Abstract
Carnation mottle virus (CarMV) is an important pathogen affecting carnation production in temperate regions.
One-step real-time RT-PCR methods using the TaqMan probe were described for quantitative detection of CarMV in carnation tissues in this paper.
Primers and probe for specific detection of CarMV were designed within the conserved region identified within the coat protein (CP) gene sequence.
A DNA fragment was amplified for mimicking the complementary RNA by PCR-based gene assembly and was used for generation of standard RNA templates.
A sensitivity assay showed that the detection limit of the assay was 200 copies, and the standard curve had a linear range from 200 to 2×106 copies, with good reproducibility.
An internal control assay designed to target the carnation 18S rRNA gene was included in detecting CarMV in infected carnation tissues.
The results showed that more positive samples were detected by RT-qPCR assay than the enzyme-linked immunosorbent assay (ELISA) when detecting the suspected samples collected from Yunnan province.
Based on presented results, the one-step real-time RT-PCR was more sensitive than ELISA for CarMV detection.
This quantitative detection assay provides a valuable tool for diagnosis and molecular studies of CarMV biology.
One-step real-time RT-PCR methods using the TaqMan probe were described for quantitative detection of CarMV in carnation tissues in this paper.
Primers and probe for specific detection of CarMV were designed within the conserved region identified within the coat protein (CP) gene sequence.
A DNA fragment was amplified for mimicking the complementary RNA by PCR-based gene assembly and was used for generation of standard RNA templates.
A sensitivity assay showed that the detection limit of the assay was 200 copies, and the standard curve had a linear range from 200 to 2×106 copies, with good reproducibility.
An internal control assay designed to target the carnation 18S rRNA gene was included in detecting CarMV in infected carnation tissues.
The results showed that more positive samples were detected by RT-qPCR assay than the enzyme-linked immunosorbent assay (ELISA) when detecting the suspected samples collected from Yunnan province.
Based on presented results, the one-step real-time RT-PCR was more sensitive than ELISA for CarMV detection.
This quantitative detection assay provides a valuable tool for diagnosis and molecular studies of CarMV biology.
Authors
Y.P. Zhang, W.Q. Huang, F. Xu, X.M. Yang, Y.L. Jiang, L.F. Zhang, L.L. Ma
Keywords
carnation mottle virus (CarMV), one step real-time RT-PCR, quantitative assay
Online Articles (30)