Additional file 1 of Improvements in lung function following vitamin C supplementation to pregnant smokers are associated with buccal DNA methylation at 5 years of age

Additional file 1: Fig. S1 Consort diagram and EWAS overview. Out of 252 pregnant smokers randomized to vitamin C or placebo as part of our VCSIP RCT, 243 offspring were delivered and 213 were re-consented into follow-up through 5 years of age. Of 192 subjects with technically acceptable spirometry at 5 years of age, 158 had sufficient quality buccal DNA for epigenome-wide methylation analysis (EWAS; n= 80 placebo, 78 vitamin C). We first performed an EWAS for vitamin C vs. placebo, followed by EWAS for FEF25-75 and current wheeze. Fig. S2 Example of scatterplot for inspection of genotype effect. We inferred that methylation was genotype dependent if there was a methylation pattern characteristic of polymorphic sites (e.g. β-values fall into two or three levels, with gaps in between, when plotted on a continuous scale of 0–1). Fig. S3 Persistent association of PSORS1C3 with FEF25-75. Among CpGs persistent over time, we highlight the correlation of one CpG (cg11811828) measured at birth versus at 5 years of age (A) and the correlation between beta values and FEF25-75 at 5 years of age (B).Fig. S4 Longitudinal plots for COL6A5. In panel A, we show the smoothed mean and confidence interval for each treatment for cg08963132 over time. In panel B, we show the correlation between raw beta values measured at birth and beta values measured at 5 years of age. Supplemental Methods.