Cancer always presents a big problem that endangers human health. In recent years, the use of gene therapy in cancer
research has significantly increased. This study aimed to construct a non-viral, wild-type, recombinant eukaryotic
expression vector, pEGFP-N1-p53/MAR and verify its mechanism of action in cancer cells in vitro. This investigation
provides a novel strategy for p53 gene therapy via regulation of the matrix attachment region (MAR), potentially laying a
foundation for the establishment of an anticancer protein bioreactor. The p53 gene was cloned from human peripheral blood
and the MAR gene was amplified from chicken liver tissue. The recombinant eukaryotic expression vector pEGFP-N1-p53/
MAR was constructed using an E. coli self-replication system. LipofectamineTM 2000 was used as the transfection agent
to deliver the plasmid into the human hepatic carcinoma (HEP3B) cell line. We divided the groups as follows: negative
control cells without plasmid transfection, vehicle control cells transfected with the PEGFP-N1 vector, and experimental
cells transfected with the pEGFP-N1-p53/MAR vector. Cells in each well of the vehicle control and experimental groups
were transfected with 1.6 μg of plasmid and 3 μL of liposome. The cellular morphology of each group was analysed
using green fluorescence microscopy at 12, 24, 36 and 48 h. Then, statistical analysis of the apoptosis rates among
the three groups was performed using SPSS. The ultrastructures of the cells were observed via transmission electron
microscopy after transfection for 24 h. Morphological analysis showed that the cells of the experimental group were
shrunken and reduced in size and their intercellular connections had disappeared. Additionally, the apoptosis rate in the
experimental group was significantly higher than that in the control groups and the cellular microstructure showed that
heterochromatin and apoptotic bodies were found in the experimental group. In conclusion, compared with the control
groups, the pEGFP-N1-p53/MAR plasmid can effectively promote Hep3B cell apoptosis in vitro.