Open AccessIdentification and characterization of the duck enteritis virus (DEV) US2 gene
Author(s) -
Jie Gao,
Anchun Cheng,
M S Wang,
Renyong Jia,
Dekang Zhu,
S Chen,
M F Liu,
F Liu,
Qiao Yang,
Kunfeng Sun,
X Y Chen
Publication year - 2015
Publication title -
genetics and molecular research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.356
H-Index - 48
ISSN - 1676-5680
DOI - 10.4238/2015.october.28.40
Subject(s) - microbiology and biotechnology , biology , polyclonal antibodies , recombinant dna , immunofluorescence , western blot , antibody , polymerase chain reaction , virus , cytoplasm , virology , gene , biochemistry , immunology
The US2 protein has been reported to contribute to duck enteritis virus (DEV) infection; however, its kinetics and localization during infection, and whether it is a component of virion, have not been previously reported. To elucidate the function of DEV US2, US2 was amplified by polymerase chain reaction (PCR) and inserted into pET-32a(+); this was expressed, the recombinant US2 protein was purified, and a polyclonal antibody generated. In addition, the kinetics and localization of the US2 gene and protein were determined by quantitative real-time fluorescent PCR, ganciclovir (GCV), and cycloheximide (CHX) treatment, western-blot, and indirect immunofluorescence assay. The packaging of US2 into DEV virions was revealed by a protease protection assay. US2 was found to be transcribed 24 h post-infection (pi) and peaked at 72 h pi; the US2 protein was detected 48 h pi, except in the presence of GCV or CHX. US2 was packed into virions and also localized to the plasma membrane and cytoplasm in infected cells. The results showed that the DEV US2 is a late gene, and that its encoding protein could be a tegument component localized mainly in the cytoplasm. This study provides useful data for the further analysis of DEV US2, including an explanation for the genetic conservation among alphaherpesviruses.