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Expression of Human T Cell Receptor ζ Chain Gene in Insect Cell
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Abstract:
Objective: To clone human T cell receptor (TCR) ζ chain gene and express its encoding protein by baculoviral expression system in insect cells. Methods: TCR ζ chain cDNA was cloned from normal human peripheral blood mononuclear cells (PBMC) by RT-PCR and inserted into baculoviral transfer vector. This vector was co-transfected with baculovirus into insect sf 9 cells. The recombinant protein expressed was identified by SDS-PAGE and flow cytometry with mouse anti-human ζ chain monoclonal antibody. Results: Human TCR ζ chain cDNA cloned and inserted into baculoviral vector specifically expressed protein in the insect sf 9 cells, accounting for about 11% of the total protein yield in the supernatant of cell lysate. The molecular weight of the recombinant protein determined by SDS-PAGE was identical to what we anticipated. The insect cells transfected with recombinant baculovirus were demonstrated by intracellular labeling flow cytometry to express ζ chain protein. Conclusion: Human T cell receptor ζ chain gene was successfully cloned from human PBMC, and its encoding protein was highly expressed in insect cells. The TCR ζ chain protein obtained by bioengineering technique is useful for in depth biological function study.
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