Objective: To explore the effect of recombinant pEGFP-N3-APC vectors carrying various APC functional domains on the expression of beta-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 beta-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 beta-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 beta-catenin level in HT-29 cells and may be applied in the gene therapy.