Objective: We established a cell line stably expressing Mycobacterium tuberculosis secretory protein EspB in order to provide evidences for studying EspB in modulating the functions of macrophage.
Methods: The recombinant plasmid pEGFP-C1-EspB was first constructed, then RAW264.7 cell was transfected with pEGFP-C1-EspB and pEGFP-C1 by liposome respectively. After screening with a high level of G418, the macrophage cell lines that stably expressed EGFP-EspB fusion protein or EGFP were established. The gene and protein expression levels were further analyzed by RT-PCR, fluorescence microscopy and western blot.
Results: The EGFP-EspB fusion gene was integrated into the chromosome and the protein was stably expressed in the selected macrophage cell line. The macrophage cell lines that stably expressed EGFP-EspB fusion protein or EGFP were established.
Conclusion: These results gave us a tool for the future study in the effects of EspB protein in modulating the functions of macrophage and its interaction with other molecules of macrophage.