Detection of anatid herpesvirus 1 gC gene by TaqMan fluorescent quantitative real-time PCR with specific primers and probe.
BACKGROUND
Anatid herpesvirus 1 (AHV-1) is known for the difficulty of monitoring and controlling, because it has a long period of asymptomatic carrier state in waterfowls. Furthermore, as a significant essential agent for viral attachment, release, stability and virulence, gC (UL44) gene and its protein product (glycoprotein C) may play a key role in the epidemiological screening. The objectives of this study were to rapidly, sensitively, quantitatively detect gC gene of AHV-1 and provide the underlying basis for further investigating pcDNA3.1-gC DNA vaccine in infected ducks by TaqMan fluorescent quantitative real-time PCR assay (FQ-PCR) with pcDNA3.1-gC plasmid.


RESULTS
The repeatable and reproducible quantitative assay was established by the standard curve with a wide dynamic range (eight logarithmic units of concentration) and very good correlation values (1.000). This protocol was able to detect as little as 1.0 x 101 DNA copies per reaction and it was highly specific to AHV-1. The TaqMan FQ-PCR assay successfully detected the gC gene in tissue samples from pcDNA3.1-gC and AHV-1 attenuated vaccine (AHV-1 Cha) strain inoculated ducks respectively.


CONCLUSIONS
The assay offers an attractive method for the detection of AHV-1, the investigation of distribution pattern of AHV-1 in vivo and molecular epidemiological screening. Meanwhile, this method could expedite related AHV-1 and gC DNA vaccine research.
Avian Disease Research Center, College of Veterinary Medicine, Sichuan Agricultural University, Yaan 625014, China.
Qing
Q
Zou
Kunfeng
K
Sun
Anchun
A
Cheng
Mingshu
M
Wang
Chao
C
Xu
Dekang
D
Zhu
Renyong
R
Jia
Qihui
Q
Luo
Yi
Y
Zhou
Zhengli
Z
Chen
Xiaoyue
X
Chen
D000818:Animals; D001715:Bird Diseases / Q000175:diagnosis* / Q000821:virology; D017931:DNA Primers / Q000235:genetics*; D004372:Ducks; D006564:Herpesviridae / Q000235:genetics / Q000302:isolation & purification*; D006566:Herpesviridae Infections / Q000175:diagnosis / Q000662:veterinary*; D015345:Oligonucleotide Probes / Q000235:genetics*; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D014764:Viral Proteins / Q000235:genetics*
D017931:DNA Primers; D015345:Oligonucleotide Probes; D014764:Viral Proteins