[Construction and expression of eukaryotic coexpression plasmid containing human MUC1 gene and GM-CSF gene].
OBJECTIVE
To construct an eukaryotic coexpression plasmid containing the coding region of human MUC1 tandem repeats gene and GM-CSF gene and to identify its expression in COS-7 cell.


METHODS
MUC1 tandem repeats gene was obtained by synthesizing the segments. After identified by restriction endonuclease digestion analysis and DNA sequencing, MUC1 tandem repeats gene and GM-CSF gene were cloned into eukaryotic expression vector pcDNA3.1(+) to construct recombinant plasmid pcDNA3.1(+)-MUC1-GM-CSF. Then the recombinant plasmid was transfected into COS-7 cell by electroporation and the expression of target gene was detected by immunoflourescence and ELISA.


RESULTS
Restriction analysis and DNA sequencing showed that the recombinant plasmid contained the coding region of human MUC1 tandem repeats gene and GM-CSF gene. The expression of MUC1 and GM-CSF was detected.


CONCLUSIONS
The suuessful construction and expression of recombinant plasmid pcDNA3.1(+)-MUC1-GM-CSF lay a foundation for further development of DNA vaccine against breast cancer.
Yuan
Shi-fang
SF
Department of Vascular and Endocrine Surgery, Xijing Hospital, China. shifangy@fmmu.edu.cn
Shi
Chang-hong
CH
Yan
Wei
W
Yao
Qing
Q
Li
Nan-lin
NL
Wang
Ting
T
Wang
Ling
L
Zhang
Ying-qi
YQ
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D004797:Enzyme-Linked Immunosorbent Assay; D005455:Fluorescent Antibody Technique; D005822:Genetic Vectors / Q000235:genetics*; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000235:genetics*; D006801:Humans; D051379:Mice; D008807:Mice, Inbred BALB C; D018396:Mucin-1 / Q000235:genetics*; D010957:Plasmids / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics* / Q000276:immunology*
C060192:MUC1 protein, human; D018396:Mucin-1; D011993:Recombinant Fusion Proteins; D016178:Granulocyte-Macrophage Colony-Stimulating Factor