[Construction of the eukaryotic expression vector of human syk and its effect on MHC-I expression of human breast cancer cells].
OBJECTIVE
To construct the eukaryotic expression vector of human spleen tyrosine kinase (Syk) and study the effect of Syk on MHC-I expression of human breast cancer cells.


METHODS
A CDS fragment of human syk was obtained by RT-PCR from human breast cancer cells MDA-MB-468 and then the fragment was cloned into the expression vector pcDNA3.1D/V5-His-TOPO. After restriction endonuclease digestion, PCR and DNA sequencing confirmation, the recombinant hsyk expression plasmid was transfected into human breast cancer cells MDA-MB-231 by using lipofectamine protocols. After G418 selection, the cells stably expressed Syk. The expression of Syk, MHC-I and ICAM-I in the transfected cells was detected by Western blot, RT-PCR and flow cytometry (FCM).


RESULTS
The eukaryotic expression plamid pcDNA3.1D/V5-His-TOPO/hsyk was constructed and it was expressed in the human breast cancer cells MDA-MB-231. The MDA-MB-231/Syk cells highly expressed MHC-I and ICAM-I.


CONCLUSIONS
The successful construction of eukaryotic expression plamid pcDNA3.1D/V5-His-TOPO/hsyk and its expression in eukaryotic cells will be useful for further study of tumor immunity.
Shan
Bao-en
BE
Research Center, Fourth Hospital of Hebei Medical University, Shijiazhuang 050011, China. baoenshan@yahoo.com.cn
Song
Xiao-tian
XT
Qi
Yi-xin
YX
D053330:4-1BB Ligand; D000818:Animals; D001483:Base Sequence; D016466:CHO Cells; D019496:Cancer Vaccines / Q000627:therapeutic use*; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D005434:Flow Cytometry; D005822:Genetic Vectors; D006801:Humans; D007371:Interferon-gamma / Q000502:physiology; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011505:Protein-Tyrosine Kinases / Q000235:genetics / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D029589:Seminal Plasma Proteins; D000072377:Syk Kinase; D014162:Transfection
D053330:4-1BB Ligand; D019496:Cancer Vaccines; D047908:Intracellular Signaling Peptides and Proteins; D009687:Nuclear Proteins; D029589:Seminal Plasma Proteins; D007371:Interferon-gamma; D011505:Protein-Tyrosine Kinases; C000606896:SYK protein, human; D000072377:Syk Kinase