**2. Results**

## *2.1. DNA Methylation*

DNA methylation raw data of all 70 IMECS participants (described in Tables A1 and A2) were normalized using Noob normalization, and beta values of all CpG sites passing QC filtering were used to calculate mean beta for each sample, as a measure for global DNA methylation. As can be seen in Figure 1A, the mean beta value for the centenarian group is slightly lower than that of the control group, however this difference was not significant. Lack of significance in this value surprisingly shows great similarity in whole-genome methylation percentage between the groups, hinting at a juvenile methylation profile for ELLI, seeing as global DNA methylation is known to increase with age [35,46]. This similarity is continued, as expected, in the offspring group, demonstrating slightly lower average beta value compared to control as well. In Figure 1B the decrease in methylation with age is easily visible, and contradicting the increase reported by Hannum et al. [35].

**Figure 1.** Mean methylation by group. Methylation levels measured via Illumina MethylationEPIC beadchip, converted to beta values with preprocessNoob to remove background read signal through Minfi R package. Ncontrol = 28, NELLI = 24, and Noffspring = 17 (one outlier removed). Difference between groups found non-significant by one-way Kruskal–Wallis analysis. (**A**) Mean beta values per group, meancontrol = 0.596 ± 0.012, meanELLI = 0.593 ± 0.008, and meanoffspring = 0.592 ± 0.011. (**B**) Linear regression of mean beta values over age, non-significant.
