BACKGROUND: Chemokine receptor 7 (CCR7) plays a key role in launching and regulating the migration of dendritic cells (DCs) from peripheral tissueto the lymphatic system. But immature dendritic cells (imDCs)do not express CCR7. Therefore, imDCs carrying CCR7 possess great prospect for stronger immune tolerance.  
OBJECTIVE: To construct green fluorescent protein (GFP) lentiviral expression vector with mouse CCR7 gene and to observe the expression of CCR7 gene in imDCs.
METHODS: The CCR7 was amplified from the mouse thymus by RT-PCR and transfected into pCR-Blunt carrier. The CCR7 DNA fragment and IRES-GFP were cloned into lentiviral expression vector LV-Lac, to form recombinant lentivial plasmid LV-CCR7. Three plasmids of the lentivial system (recombinant lentivial plasmid LV-CCR7, package plasmid ΔNRF, envelope plasmid pVSVG) were co-transfected with package lentivirus by lipofectamine. The imDCs were infected by the recombinant lentivial, The morphology of transfected imDCs was examined under a fluorescent microscope and flow cytometry was used to determine the expression of CCR7.
RESULTS AND CONCLUSION: The CCR7 fragment was amplified from cDNA of the mouse thymus and subcloned to pCR-Blunt carrier, and lentiviral expression vector LV-CCR7 was constructed successfully. After infected with 293 FT cells for 24 hours through three plasmid package system, positive expression of GFP was observed under fluorescence microscope. The titer of the lentivirus was above 108 U/L, and to obtain the recombinant lentivial with mouse CCR7 gene. The lentivial plasmid could infect imDCs effectively, a large amount of GFP and CCR7 expression was observed under fluorescence microscope and flow cytometry, respectively, and the positive rate was 50%. It indicates that GFP lentiviral vector LV-CCR7 with mouse CCR7 gene were successfully constructed and expressed in imDCs.