© 2012 Landes Bioscience. Do not distribute. Figure 4. Distance to transcription start site (Tss ) influences methylation of probes in promoters. 98% of probes on the Illumina Infinium h umanMeth- ylation27 Beadc hip array were within 1,500 bps of a known gene Tss . (a ) In the merged villi group, DNa methylation was analyzed by binning probes into six 500 bp windows around known Tss . There were significant differences in DNa methylation between probes in strong (●) weak (■) and non (▲) islands (p < 0.0001) but direction to Tss had no significant effect on DNa methylation. DNa methylation of probes furthest from Tss (±1000 to 1500 bps; 16.31% ± 2.26, 59.29% ± 5.39, 63.82% ± 3.6 for strong, weak and non islands respectively) was significantly higher than probes close to Tss (±0 to 500 bps; 8.65% ± 0.77, 35.60% ± 2.16, 57.14 ± 3.33 for strong, weak and non islands respectively, all p<0.001). Tissue differences in DNa methylation of 2nd trimester villi, muscle, kidney, brain and adult blood were investigated in three Tss bins in strong c pG islands: (B) ±0 to 500 bps, (c ) ±501 to 1,000 bps and (D) ±1,000 to 1,500 bps. s ignificance is indicated by *p < 0.05, **p < 0.001, ***p < 0.0001. with r > 0.5 than for r < -0.5 (Fig. S12; 8 vs. 1). These results sug- gest a general trend for correlation of DNA methylation between tissues; however, conclusions are limited by the small sample size. Increase of villi weak and non-island probe methylation throughout gestation. We previously reported an increase in villi gene promoter methylation throughout gestation using the Illumina Infinium HumanMethylation27 BeadChip array data for 1st trimester, 2nd trimester and term villi.35 Here, we evaluated gestational age changes in villi DNA methylation with the addition of LINE-1 and Alu DNA methylation, and further subdivision of array probes into strong, weak and non-islands (Fig. 5A–E). After correction for multiple comparisons, there was a significant increase in non-island probe methylation from 1st trimester (46.99% ± 1.75) to term (52.01% ± 0.91) (Fig. 5E; p < 0.0001) and weak island probe methylation from 1st trimester (36.12% ± 0.99) to term (38.10% ± 0.68) (Fig. 5D; p < 0.0001) 658 Epigenetics Volume 7 Issue 6