Identification of mESC Raman signatures due to an altered global DNA methylation. (a) Raman spectra of mESCs cultured in (i) control and (ii) 2i medium. Raman signatures that can be assigned to DNA bands are indicated by black arrows. The standard deviation is indicated by the gray lines. (b) Statistical analysis of significantly increased Raman bands, which can be assigned to DNA methylation. Two-tailed t-test, n = 85. (c) Score plot of PC 1 and PC 3 shows a grouping tendency of mESCs cultured in control and 2i media. The direction of the gray arrow indicates the loss of DNA methylation. The score mean value of both groups for PC 4 differs significantly from each other. Two-tailed t-test, n = 85 (d) PC 4 loading describes the Raman shifts that vary in 2i-adapted mESCs compared with mESCs cultured in the control medium. Peaks of interest are indicated by black arrows.