Methylome and transcriptome sequencing to study epigenetic alterations in colon associated fibroblasts from normal, colitis and cancer associated fibroblasts

The colon cancer associated fibroblasts are part of the tumor microenvironment and has been shown to facilitate colon cancer progression by secretion of various signaling molecules and growth hormones. Similarly, the fibroblasts associated with colitic colons secrete pro-inflammatory cytokines that promote inflammation of the colon. However, the molecular mechanisms that underlie these gene expression changes in these fibroblasts is unclear. To characterize the epigenetic mechanisms that may contribute to the gene expression changes in these fibroblasts, we performed RNA-seq and MBD-isolated genome sequencing (MiGS or MBD-seq) on colon associated fibroblasts isolated from normal, colitis and cancer patients. DNA and RNA were extracted from cultured colon associated fibroblasts from normal, colitis and cancer patients using Qiagen Allprep Kit. The DNA samples were used for MBD-isolated genome sequencing (MiGS), followed by addition of illumina adapters for high-throughput sequencing. The RNA samples were used for paired end Stranded RNA-seq.