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Retroviral Overexpression of CXCR4 on Murine B-1a Cells and Adoptive Transfer for Targeted B-1a Cell Migration to the Bone Marrow and IgM Production
Published on: May 31, 2020
[Construction of human CXCR3B gene transfected cell line, identification and study of its biological function]
Jing-ya Guo1, Chang-you Chen, Zhi-qiang Wang
1Department of Immunology, Medical College of Soochow University, Suzhou 215123, China. guojingya1118@163.com
Aim:
To construct recombinant human CXCR3B gene expression vector and obtain L929-CXCR3B gene transfected cell line for stably expressing human CXCR3B.
Methods:
Human CXCR3B gene of full length was amplified by PCR from the plasmid pMD19-T/huCXCR3A. Then, it was inserted into eukaryotic expression vector pIRES2-EGFP to construct recombinant vector pIRES2-EGFP/huCXCR3B. The recombinant vector was transfected into L929 cells with Lipofectamine(TM); 2000, and cell clones stably expressing human CXCR3B molecule were screened by G418.We used FCM and RT-PCR to verify expression of CXCR3B from protein level and gene level. The ability of proliferation of L929-huCXCR3B under the circumstance of CXCR3B ligand called Mig was analyzed via MTT methods.
Results:
We have constructed recombinant human CXCR3B gene expression vector and obtained L929-huCXCR3B gene transfected cell line which can stably express human CXCR3B molecule. The positive expression rate of CXCR3B on L929-huCXCR3B cells was 93&. The result of MTT assay showed that the proliferation of L929-huCXCR3B cells were inhibited when the cells were cocultured with Mig after 24 h, 48 h and 72 h, and the inhibition ratio were 41.44&, 44.01& and 24.80& respectively.
Conclusion:
Construction of L929-huCXCR3B cell line has laid a good foundation on research of the CXCR3B signal pathway and preparation of the CXCR3B monoclonal antibody.
