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Published on: May 18, 2020
[Construction of PTEN eukaryotic expression plasmid and its effects on breast carcinoma cell line MDA468]
Qing-yong Chen1, Dao-da Chen, Chun-you Wang
1Department of General Surgery, Xiehe Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China. chenqy11@hotmail.com
Objective:
To investigate the effects of exogenous wild PTEN gene stably transfection on growth of breast cancer cells in vitro.
Methods:
At first, a recombinant eukaryotic expression plasmid pcDNA3.1-PTEN was constructed. Human breast cancer cell line MDA468 was transfected with pcDNA3.1-PTEN or mock transfected plasmid pcDNA3.1(-) with lipofectamine. RT-PCR, immunohistochemical staining and Western blot were used to determine target gene expression. Cell viability was tested by MTT assay. Apoptosis was determined by flow cytometry with a double-staining method using FITC-conjugated annexin V and PI.
Results:
The PTEN stably transfected cells demonstrated the integration of the exogenous target gene and corresponding mRNA and protein over-expression. There was a significant decline in cell viability of pcDNA3.1-PTEN transfected MDA468 cells in comparison with the mock-transfected ones (P < 0.01). The PTEN-trasfected MDA468 cells also showed an increase in the rate of apoptosis, compared with parental and mock-trasfected cells (P < 0.001).
Conclusion:
Stable expression of exogenous PTEN can suppress the malignant phenotypes of the human breast cancer cell line MDA468.
