*5.6. ChIP Assay*

22Rv1 cells were incubated in the presence or absence of 50 μM NCL1 as indicated. Formaldehyde (1%) was used to cross-link cells, and the chromatin was collected and subjected to immunoprecipitation using an H3K4-me2 antibody (Cell Signaling Technology, Beverly, MA, USA). As a negative control, isotype-specific IgG was used. Extracted DNA was dissolved in TE buffer, and real-time PCR using specific primers (F-GGGGCGGTTGTATATCAGG, R-GGCTCCACAAGGAACTGACT) was used to confirm the methylation status of the promoter regions of P21 (CDKN1A).
