Figure 2

Bisulfite sequencing of DNA CpG rich region in primary tissue samples from patients with CRC in the proximal (A) and distal (B) colon. The primary cancerous and histopathologically unchanged tissues from the same patients with cancer in the proximal (P1-P5) and distal colon (P6-P10) were used for genomic DNA isolation followed by bisulfite conversion of cytosine to uracil. The HSD17B1 region containing 31 CpG dinucleotides (chr17: 37 953 392-37 953 917) was then amplified by a pair of primers complementary to the bisulfite-DNA modified sequence (Additional file 1, Additional file 2). The PCR products were purified with subsequent cloning into a plasmid vector. Plasmid DNA isolated from ten positive bacterial clones was used for commercial sequencing. The results of bisulfite sequencing were assessed and presented using BiQ analyzer software and BDPC web server, respectively [22, 23]. Black and grey boxes represent methylated and unmethylated CpG dinucleotide, respectively. White boxes correspond to undetermined CpG dinucleotide. The legend with grey scale corresponds to average methylation in (P1-P5) and (P6-P10).