Figure 4

Comparison of MethyLight and Matrix ChIP-MeDIP assay of normal cervix tissue and cervical cancer. MethyLight, DNA purified from clinical cancer and normal tissues was bisulfite-converted and amplified using SPARC gene primer in Taqman PCR [43]. Methylight results are expressed as PMR values for each individual sample. ChIP-MeDIP, frozen (-80°C) archived cervical tissue samples histologically classified as normal and cancer were thawed, minced and fixed with formaldehyde. After glycine treatment and washes samples were treated with high energy ultrasound to shear the chromatin. The lysates were cleared by centrifugation, aliquoted and stored (-80°C). For MeDIP the input was deproteinized, RNase-treated and denatured DNA. MeDIP was done using anti-5mC antibody (Aviva) and ChIP assays with anti-Pol II CTD, H3K9/14Ac, H2A.Z and H3K27m3 antibodies. MeDIP and ChIP DNA were analyzed at the indicated site of the SPARC gene in real-time PCR. Data represent values for each individual sample (five normal and five cancer), expressed as % input.