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Development of a real-time fluorescent quantitative PCR assay for detection of Impatiens necrotic spot virus.

Abstract：

:Impatiens necrotic spot virus (INSV) is an important plant virus that can cause severe disease in various ornamental and agricultural crops. Several species of thrips transmit INSV, of which the western flower thrip (Frankliniella occidentalis) is the most important. In this study, primers and TaqMan probes based on INSV non-structural protein gene sequences were designed, and a technique was developed for detecting INSV using fluorescent quantitative RT-PCR. The reproducibility, specificity and sensitivity for the RT-PCR were evaluated; and the RT-PCR method was developed to detect INSV in the host plants and western flower thrips. A standard curve constructed by a series of diluted plasmid DNA gave a good linear relationship between Ct value and concentration of plasmid DNA, a low coefficient of variation and good reproducibility. The detection method not only measured quantitatively the concentration of INSV in plant hosts and western flower thrips, but also measured accurately low concentrations of the virus. The measurable concentration fell to as low as 10⁰ copies/μl, while RT-PCR could detect only 10² copies/μl. The method had high specificity and could distinguish INSV from Tomato spotted wilt virus (TSWV) and Tomato zonate spot virus (TZSV), both from the same genus of viruses. This is the first report of the same method being used to detect INSV in both plant hosts and western flower thrips, and should be helpful in studies of INSV epidemiology.

journal_name

J Virol Methods

journal_title

Journal of virological methods

authors

Chen X,Xu X,Li Y,Liu Y

doi

10.1016/j.jviromet.2013.02.012

subject

Has Abstract

pub_date

2013-05-01 00:00:00

pages

299-304

issue

2

eissn

0166-0934

issn

1879-0984

pii

S0166-0934(13)00061-X

journal_volume

189

pub_type

杂志文章

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