Effect of recombinant pEGFP-N3-APC vectors carrying various APC functional domains on the expression of β-catenin in HT-29 cells / 中南大学学报(医学版)

ABSTRACT

Objective To explore the effect of recombinant pEGFP-N3-APC vectors carrying various APC functional domains on the expression of β-catenin in human colorectal cancer cells HT-29.Methods The recombinant plasmids were transfected into HT-29 cells mediated by lipofectamine~(TM) 2000, and detected by green fluorescence and RT-PCR. Western blot was applied to detect β-catenin expression level in HT-29 cells after transfection, and gray scales of electrophoresis strips were analyzed by SPSS 13.0.Results Green fluorescence and RT-PCR made clear that all 5 recombinant plasmids were successfully expressed in HT-29 cells. Western blot showed that β-catenin expression level in HT-29 cells was not affected after being transfected with pEGFP-N3-APC1, pEGFP-N3-APC2 and pEGFP-N3-APC3, and was distinctly affected after being transfected with pEGFP-N3-APC4 and pEGFP-N3-APC5, especially the later one. Conclusion The selected APC5 gene fragment with 15-amino acid repeats and SAMP repeats, which is relatively short, can degrade β-catenin level in HT-29 cells and may be applied in the gene therapy.