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  • Title: [Demethylation in the promoter area by the antisense of human DNA MTase gene].
    Author: Zhang H, Xiao W, Liang H, Fang D, Yang S, Luo Y.
    Journal: Zhonghua Zhong Liu Za Zhi; 2002 Sep; 24(5):444-7. PubMed ID: 12485494.
    Abstract:
    OBJECTIVE: To investigate the change in the expression of E-Cadherin of human hepatocarcinoma cell line SMMC-7721 after transfection by antisense human DNA MTase gene. METHODS: DNA MTase gene eukaryotic expression vectors, including sense and antisense fragments, were constructed with recombinant technology and transfected into the hepatocarcinoma cell line SMMC-7721 with liposome DOTAP. The expression of DNA MTase gene mRNA and E-Cadherin gene mRNA was examined with RT-PCR and the expression of E-Cadherin with immunohistochemical and flow cytometry. The status of methylation in E-Cadherin gene promoter area was examined with methylation specific PCR (MSP). RESULTS: The sense and antisense eukaryotic expression vectors were successfully constructed and then the constructed recombinant plasmids were successfully transfected into SMMC-7721 cell with liposome DOTAP. The expression of endogenous DNA MTase mRNA was obviously decreased with E-Cadherin gene mRNA and its activity increased in the SMMC-7721 cell, which was tranfected with antisense DNA MTase gene fragment. Moreover, demethylation in the promoter area of E-Cadherin gene was observed with MSP. CONCLUSION: Demethylation in the promoter area and increasing mRNA level of E-Cadherin gene can be induced by expression inhibition of DNA MTase gene of SMMC-7721 cell line.
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