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Context DNA methylation in the diagnosis of gestational diabetes Objective To assess the value of DNA methylation in the diagnosis of gestational diabetes (GDM) and in the prediction of maternal postpartum glucose disturbances. Design Two-stage observational between July 2006 and December 2010. Setting University Hospital Patients Forty-eight randomly selected pregnant women formed the discovery cohort (24 with GDM and 24 controls) and 252 pregnant women (94 with GDM and 158 controls) formed the replication cohort. GDM women were re-evaluated 4 years postpartum. Main Outcome Measures GDM, type 2 diabetes or prediabetes at 4 years postpartum Results We identified three CpG sites related to LINC00917, TRAPPC9 and LEF1 that were differentially methylated in women with GDM and abnormal glucose tolerance, and the sites associated with LINC00917 and TRAPPC9 were independently associated with an abnormal glucose tolerance status four-years postpartum after controlling for clinical variables. Moreover, the site associated with LINC00917 and the combination of the three sites had the highest predictive values. Conclusions Our results suggest that some of these sites may be implicated in the development of GDM and postpartum abnormal glucose tolerance.