[Effect of recombinant pEGFP-N3-APC vectors carrying various APC functional domains on the expression of beta-catenin in HT-29 cells].
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.


CONCLUSIONS
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.
Lu
Liang
L
Department of Gastroenterology, Second Xiangya Hospital, Central South University, Changsha 430011, China.
Huo
Jirong
J
Liu
Jia
J
Wu
Jie
J
Wang
Jie
J
D025601:Adenomatous Polyposis Coli Protein / Q000096:biosynthesis / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000096:biosynthesis / Q000235:genetics*; D019073:HT29 Cells; D006801:Humans; D017434:Protein Structure, Tertiary / Q000235:genetics; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection; D051176:beta Catenin / Q000096:biosynthesis* / Q000235:genetics
C555337:APC protein, human; D025601:Adenomatous Polyposis Coli Protein; D011994:Recombinant Proteins; D051176:beta Catenin; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins