The 6th amino acid mutation of Rep protein had no
effect on PCV2b but enhanced PCV2d virus replication in vitro
Xiaoyan Wu1,2,a, Shuo
Wang2,a, Changxun
Xin 2,a,
Chen Li2, Jianli Shi2, Zhe
Peng2, Chang Liu2, Hong
Han2, YaoTian3,
JiaxinLi3, Shaojian Xu2, Jun
Li1,2,3*,Fan Zhang1*
1 Shandong Normal University, Jinan, 250014, China.
2Division of Swine Diseases, Shandong Provincial Key Laboratory of Animal
Disease Control & Breeding, Institute of
Animal Science and Veterinary
Medicine Shandong Academy of
Agricultural Sciences, Jinan, 250100, China
3 Qingdao Agricultural University, Qingdao, 266000,
China.
* Address for correspondence: Jun Li, Shandong Provincial Key Laboratory
of Animal Disease Control & Breeding,
Institute of Animal Science and
Veterinary Medicine Shandong Academy of Agricultural
Sciences,
No.202, North Industrial Rd., Jinan 250100, People’s Republic of China;
E-mail: junli79@163.com.
Fan Zhang, Key Laboratory of Animal Resistance Research, College of Life
Science, Shandong Normal University, 88 East Wenhua Rd, Jinan 250014,
People’s Republic of China; E-mail:
zhangfan0531@163.com
a These authors contributed equally to this work.
Abstract: Porcine circovirus type 2 (PCV2) is the etiological
agent that primary cause of post-weaning multisystemic wasting syndrome
(PMWS). The major genotypes, PCV2a, PCV2b and PCV2d, are highly
prevalent, but now replaced with 2b and 2d in swine population in
worldwide. Rep protein is the key protein for viral replication.
Compared a large number of Rep protein amino acid (aa) sequences, we
found that there were three sites with regular changes between 2b and
2d.In order to analyze the effect of key sites on viral replication, we
used site-directed mutagenesis to mutate the 6th aa of
Rep (alternations with asparagine and serine) between PCV2b and PCV2d,
Two wild-type and two mutant viruses infectious clones were rescued by
non-contaminated porcine kidney-15 (PK-15) cells. Real-time quantitative
PCR and a one-step growth curve were used to determine viral load to
assess the replication of rescued viruses. The results showed that there
was no significant difference
between
the PCV2b mutation and the wild-type PCV2b virus in vitro, while the
mutation of PCV2d enhanced viral replication.
Keywords: Porcine circovirus type 2; Rep protein; mutate sites;
viral replication