Construction And Analysis Of The Characteristic Of Canine Parvovirus Virus-like Particles Coated With Quantum Dot Composites

Virus-like particles (VLPs) of Canine parvovirus (CPV) are polymer particles which have a similar spatial structure with natural virus particles that consist of canine parvovirus capsid protein. Because Canine parvovirus VLPs do not contain any virus genetic material and have good immunogenicity and safety, they can effectively stimulate the body to produce a good immune response. In recent years, the use of VLPs obtained from CPV VP2 protein show that they not only maintain the immunogenicity of the VLP but also simulate the infection pathway of natural virus particles of CPV. VP2 protein can achieve targeting position of CPV VLPs by specific cell surface receptor binding. Quantum dots (QD) with its unique fluorescence properties have good potential in biological imaging, but the poor biocompatibility, no ability of target binding, and the obvious toxicity to host cell and body, severely limit the application of the quantum dots in biology.In this study, we used the PCR method to amplify the CPV VP2 capsid protein gene. Afterwards, linked this gene to the pSMK plasmid. By soluble E. coli expression system, we expressed the soluble protein VP2, then used the viral capsid protein to self-assemble into virus-like particle. In vitro environment, took the MPA modified QDs MPA-QDs into VLPs to form a fluorescent compound which had good biocompatibility, and no biological toxicity.1. This research verified the modified QDs by MAA and MPA. Nanosizer revealed that after being modified, surface charge of MAA-QD and MPA-QD became to be negative charge compared with the unmidified QDs which neutrally charged. And this negative charge was also been confirmed by nucleic acid electrophoresis.2. We structured the expression plasmid pSMK-VP2, and came true the epression of the protein HIS-SUMO-VP2 in Escherichia coli expression system. Chromatographic purification on Ni-NTA was performed and followed by SUMO protease digestion to cut off the His-SUMO on the N-terminal. Then we got the VLPs that recognized by CPV monoclonal antibody. After the VLPs coated the MPA modofied QDs, we finally got the CPV VLP-QDs. By means of the TEM, nucleic acid electrophoresis, ultraviolet fluorescence properties, and nano diameter, this CPV VLP-QDs complex was verified to be better biocompatibility, lower cytotoxicity, better fluorescent brightness, better environmental stability.This research successfully structured the CPV VLP-QDs complex with etter biocompatibility, lower cytotoxicity, better fluorescent brightness, better environmental stability, which provides a basic research for study on the invasion by CPV virions of the cells, and their location in the host cells. The follow-up study could be immunization of this CPV VLP-QDs complex, by observing the fluorescence in cell or between organizations, could we provide a basic research on mechanism of immunization caused by VLPs.