The DNA methylation status of the hCD2 transgene in hCD2+ and hCD2- T cells from the CD2 1.3 transgenic lines. (A) Schematic transgene map showing the restriction sites for Bgl II (B) and Hha I (H) and location of the probe used in B. (B) The CpG methylation analysis of the proximal promoter region by methylation-sensitive restriction enzyme digest and Southern blot. The bar charts show the ratios between the unmethylated and methylated Hha I sites. (C) Bisulfite sequencing analysis of the 3' regulatory regions of the hCD2 transgene. Methylated and unmethylated CpGs are shown as filled and open circles, respectively. Each line represents the sequence from a single clone. Similar results were obtained from two independent experiments. Comparisons between numbers of methylated CpGs were done using the Mann-Whitney U-test and Fisher's exact test (see text).