Figure 2
From: A CRISPR-based approach for targeted DNA demethylation
RANKL gene upregulation showed DNA demethylation-dependent manner. (a) Quantitative real-time PCR (qRT-PCR) assay was performed to assess the abundances of RANKL mRNA in HEK-293FT cells in indicated groups. (b) The percentages of methylated or unmethylated DNA at each site in RANKL gene, determined by bisulfite sequencing, were represented by green or white sections in the filled circles, respectively, with the numeric information listed in Supplementary Table S2. The sgRNAs recognizing their respective target sites were shown in blue-pink color (as in Figure 1e). (c) mRNA levels of RANKL were examined in HEK-293FT cell that stably expressed the demethylation system using qRT-PCR assay. (d) In the HEK-293FT cells stably expressing the demethylation system, the percentages of methylated or unmethylated DNA at each site in RANKL gene were determined through bisulfite sequencing, respectively. The corresponding detailed numeric data were listed in Supplementary Table S3. The sgRNAs recognizing their respective target sites were shown in blue-pink color (as in Figure 1e). (e) RANKL mRNA expression was assayed 4 days after co-transfection of sgRANKL-(1-8) guided dCas9- and MS2-Tet1-CD in SH-SY5Y cell line using qRT-PCR assay. Data were shown after normalization to the controls (blank group) (means±s.e.m., n=3). *P<0.05, compared with -sgRNA group by unpaired t-test.
