[Construction of the eukaryotic expression vector of human syk and its effect on MHC-I expression of human breast cancer cells].

Aim: 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.

Conclusion: 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.