Fig 1: Bisulfite amplicon sanger sequencing.Standard curves were generated by sanger methylation quantification of known methylation level controls. The expected percentage of methylation was plotted versus the observed one. Points represent the mean methylation level of the fifteen and eleven CpG sites for the two regions, respectively, as analyzed with Sanger/ESME (blue line = Lambda control 1, red line = Lambda control 2). Both ?-DNA controls were able to fit linear lines with R2 values as presented. The correlation coefficients are higher in the data generated from the Premium Bisulfite kit (Diagenode) and the MethylEdge Bisulfite Conversion System (Promega). [P value for i) Lambda control 2 (Qiagen) = 0.1309, ii) rest of samples < 0.05].
Fig 2: Bisulfite conversion efficiency of the different kits.The conversion efficiency of the unmethylated cytosines to thymines was calculated per kit. All the non-CpG cytosines of all the 24 libraries of the Lambda control 1 were used for the calculation. Conversion efficiency of the Premium Bisulfite kit (Diagenode) was 99.0±0.035%, of the MethylEdge Bisulfite Conversion System (Promega) was 99.8±0.000%, of the EpiTect Bisulfite kit (Qiagen) was 98.4±0.013% and of the BisulFlash DNA Modification kit (Epigentek) was 97.9%. Error bars = SD.
Supplier Page from Promega for MethylEdge Bisulfite Conversion System