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. 2021 Nov 8;47(1):10. doi: 10.3892/or.2021.8221

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Copyright: © Frazzi et al.

This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivs License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non-commercial and no modifications or adaptations are made.

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Figure 3. — Effects of DNMT1- and DNMT3a-selective silencing on gene-specific quantitative methylation and expression. (A) Silencing efficiency after 48 h from transfection in Toledo and NU-DUL-1 cell lines as determined by RT-qPCR. *P<0.05, **P<0.01, ***P<0.0001 (n=3; one-way ANOVA). (B) Silencing efficiency 48 h after transfection in Toledo and NU-DUL-1 cell lines as determined by immunoblotting (n=2). (C) Gene expression levels of KLF4, DAPK1 and SPG20 as determined by RT-qPCR (n=2). (D) Quantitative promoter methylation in the panel of target loci 48 h after DNMT1 silencing. (E) Quantitative promoter methylation in the panel of target loci 48 h after DNMT3a silencing. (F) Quantitative promoter methylation in the panel of target loci 48 h after DNMT1 + DNMT3a silencing (n=2/3). (G) Ki67 staining of the transfected cells after 48 h incubation as determined via immunocytochemistry (magnification, ×40). (H) Cell Counting Kit-8 viability assay of the transfected cells after 48 h incubation. *P<0.05 (one-way ANOVA). DNMT, DNA methyltransferase; RT-qPCR, reverse transcription-quantitative PCR; KLF4, Krüppel-like factor 4; DAPK1, death associated protein 1; SPG20, spastic paraplegia 20; si, small interfering RNA.