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Establishment and evaluation of real-time PCR for West Nile virus detection

Published online by Cambridge University Press:  24 April 2009

Shi Li-Jun ,
Lu Mao-Min ,
Li Gang ,
Li Cheng-Yao  and
Zhang Jin-Gang
Shi Li-Jun
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China Institute of Animal and Veterinary Medicine, Chinese Academy of Agricultural Science, Beijing 100193, China
Lu Mao-Min
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China
Li Gang
Affiliation:
Institute of Animal and Veterinary Medicine, Chinese Academy of Agricultural Science, Beijing 100193, China
Li Cheng-Yao
Affiliation:
College of Bio-technology, Southern Medical University, Guangzhou 510515, China
Zhang Jin-Gang*
Affiliation:
Institute of Transfusion Medicine, Academy of Military Medical Sciences, Beijing 100850, China
*
*Corresponding author. E-mail: [email protected]
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Abstract

A rapid real-time polymerase chain reaction (RT-PCR) for detecting West Nile virus (WNV) was established. Primers were designed according to the sequence of the capsid protein gene of WNV by Primer Premier 5.0. In this way, an inexpensive assay using the intercalating dye SYBR Green I was developed and validated. The amplifying curve showed that this method could successfully amplify 102 copies/μl of the WNV gene, while reference to Japanese encephalitis virus (JEV) and blank control were all negative. Tenfold successive dilutions of positive WNV DNA were used to measure the sensitivity of RT-PCR. The assay system showed high reproducibility with coefficient of variation (CV) <2%. Thus the newly established RT-PCR assay was shown to be a rapid, sensitive and specific test for detecting WNV.

Keywords

West Nile viruscapsid protein genereal-time PCR
Type
Research Papers
Information
Chinese Journal of Agricultural Biotechnology , Volume 6 , Issue 1 , April 2009 , pp. 55 - 59
Copyright
Copyright © China Agricultural University 2009

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