Abstract:
Background Hematopoietic pre-B cell leukemia transcription factor (PBX)-interacting protein (HPIP) is involved in the development of many malignant tumors, but its mechanism in lung cancer remains to be explored.
Objective To construct the eukaryotic expression vector of HPIP labeled with Flag-tag, express the pcDNA3.0-Flag-HPIP fusion protein, and detect its effect on the proliferation, migration, and other biological functions of human lung cancer cells.
Methods Using human breast library as template, the coding sequence of HPIP gene was amplified by PCR technology, and inserted into pcDNA3.0-Flag vector. The recombinant plasmid was transfected into A549 lung cancer cell line, and the expression of HPIP after transfection was verified by western blotting. The effect of the protein on the cell proliferation and migration of lung cancer cell line were assessed by CCK-8, clone formation assay and wound healing assay, respectively.
Results The encoding sequence of HPIP gene with a length of about 2 109 bp was amplified by PCR and successfully inserted into pcDNA3.0-Flag vector. Sequencing and western blotting confirmed the successful construction of the fusion protein. CCK-8 assay and clone formation assay showed that the proliferation ability of lung cancer A549 cells transfected with Flag-HPIP was significantly enhanced compared with the control group. Wound healing assay confirmed that the transfection of Flag-HPIP could significantly improve the migration ability of lung cancer A549 cells.
Conclusion The eukaryotic expression vector of HPIP gene with Flag label is successfully constructed, and it is confirmed that the fusion protein can promote the proliferation and migration of lung cancer cells, which lays a good foundation for the subsequent functional studies of HPIP in lung cancer.
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