[Secreting expression and characterization of canine parvovirus VP2 protein in eukaryotic cells].
OBJECTIVE
To study and characterize secretive expression of canine parvovirus capsid protein 2 (VP2) gene in eukaryotic cells.


METHODS
To construct secreting expression vector of VP2 gene, we obtained CD5 signal peptide (SP) DNA fragment from plasmid containing human CD5 SP DNA sequence and inserted the fragment into multiple clone site of eukaryotic expression vector pcDNA3.1A. The canine parvovirus VP2 gene was amplified by PCR and inserted into expression vector pcDNA3.1-CD5sp down stream of CD5 SP. The recombinant pcDNA-CD5sp-VP2 plasmids were transfected into HEK293T cells mediated by calcium phosphate. VP2 binding activity for canine transferrin receptor was analyzed by ELISA method.


RESULTS
Recombinant pcDNA-CD5sp-VP2 plasmid proved to be correct by sequencing. VP2 proteins were detected by Western-blot in the culture medium of transfected 293T cells, which indicated that the expressed VP2 protein could be secreted into the medium mediated by human CD5 SP. VP2 protein had the activity to bind canine transferrin receptor (TfR).


CONCLUSIONS
The secreting expression of VP2 in eukaryotic cells was achieved by using human CD5 SP. Recombinant VP2 showed the ability to bind canine TfR.
Wang
Wei
W
Department of Basic Veterinary Medicine, College of Animal Science and Technology, Agricultural University of Hebei, Baoding 071001, China. airenwu@126.com
Li
Xiujin
X
Zhong
Fei
F
Wang
Xingxing
X
Han
Dongmei
D
Jin
Huijun
H
Pan
Sumin
S
Li
Wei
W
D000818:Animals; D036022:Capsid Proteins / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D006224:Cricetinae; D004285:Dogs; D004797:Enzyme-Linked Immunosorbent Assay; D005057:Eukaryotic Cells / Q000378:metabolism; D005822:Genetic Vectors; D006801:Humans; D017993:Parvovirus, Canine / Q000737:chemistry / Q000235:genetics*; D016133:Polymerase Chain Reaction; D011990:Receptors, Transferrin / Q000378:metabolism; D014162:Transfection
D036022:Capsid Proteins; D011990:Receptors, Transferrin; C095548:capsid protein VP2, parvovirus B19