Targeted bisulfite sequencing of the H19 imprinting control region and the LINE1 repetitive elements was performed to estimate maintenance methylation of Dnmt1-deficient intestinal crypt cells. Crypt epithelium was isolated by laser capture microdissection from controls and mutants at one week (n=3–4 per group), one month (n=2 for controls, n=4 for mutants), or two months (n=4–7 per group) following tamoxifen treatment.
(A,C,E) Targeted bisulfite sequencing of six CpGs in the H19 imprinting control region (Chr7: 149,766,621-149,766,690). At one week (A), one month (C) and two months (E) following tamoxifen administration, H19 methylation levels are comparable between Dnmt1 mutants and controls.
(B,D,F) Targeted bisulfite sequencing of nine CpGs in the LINE1 repetitive elements (NCBI Accession #D84391: 976-1,072). LINE1 methylation levels one week after tamoxifen injection are significantly reduced in crypts of mutant mice compared to controls (B). One month (D) and two months (F) after tamoxifen-induced ablation of Dnmt1, global DNA methylation levels have been restored to baseline.
For all graphs, data are presented as average±SEM. **P<0.01 by two-tailed Student’s t-test.