[Construction and expression of fusion gene expression plasmid HBV preS2S-rhGM-CSF].
OBJECTIVE
To construct GM-CSF and preS2 fusion gene expression plasmid to enhance the immunogenicity of hepatitis B DNA vaccine.


METHODS
HBV preS2S gene(846 bp) and rhGM-CSF gene(384 bp) were amplified by PCR, respectively. The eukaryotic expression plasmid pcDNA3.1-S2S-rhGM-CSF was constructed by means of T-A clone and directional gene cloning techniques, and then the recombinant plasmid was expressed in HepG2 cells.


RESULTS
Restriction enzyme digestion analysis, PCR amplification and/or DNA sequencing proved that the recombinant plasmid was constructed successfully. The transcription of target gene was confirmed by RT-PCR. The fusion protein expressed in HepG2 cells could reacted to the monoclonal antibodies (mAbs) against HBsAg, preS2 and GM-CSF, respectively.


CONCLUSIONS
The successful construction and expression of pcDNA3.1-S2S-rhGM-CSF lay the foundation for further study of HBV DNA vaccine.
Guo
Xiao-lan
XL
Department of Clinical Laboratory, Affiliated Hospital of North Sichuan Medical College, Nanchong 637000, China.
Deng
Jian-kang
JK
Zhu
Dao-yin
DY
Tang
En-jie
EJ
D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D045744:Cell Line, Tumor; D015967:Gene Expression Regulation, Viral; D005814:Genes, Viral; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D006514:Hepatitis B Surface Antigens / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D006515:Hepatitis B virus / Q000235:genetics* / Q000276:immunology; D018197:Hepatoblastoma / Q000378:metabolism / Q000473:pathology; D006801:Humans; D008113:Liver Neoplasms / Q000378:metabolism* / Q000473:pathology; D008969:Molecular Sequence Data; D010957:Plasmids; D011498:Protein Precursors / Q000235:genetics* / Q000276:immunology / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000378:metabolism; D014162:Transfection
D000911:Antibodies, Monoclonal; D006514:Hepatitis B Surface Antigens; D011498:Protein Precursors; D011993:Recombinant Fusion Proteins; C050807:presurface protein 2, hepatitis B surface antigen; D016178:Granulocyte-Macrophage Colony-Stimulating Factor