DNA-methylation analysis of 5′ CpG islands at the GPC3 and SYBL1 loci. Genomic DNA was used in Southern blotting at the GPC3 (A) and SYBL1 (B) loci. (A) Genomic DNA from Raji cells (lanes 1, 5, and 9), Daudi cells (lanes 2, 6, and 10), THX88 (inactive X chromosome-containing hybrid cell line) cells (lanes 3, 7, and 11), and Caco-2 cells (lanes 4, 8, and 12) was restricted with HindIII in combination with KspI (lanes 1–4), EclXI (lanes 5–8), or BssHII (lanes 9–12), and Southern blotting was performed by using a 1.4-kb NheI fragment encompassing the GPC3 CpG island as the probe. (B) This Southern blot is similar to the one shown in A, but it uses the SYBL1 1.6-kb EcoRI fragment as the probe with normal male (lanes 1 and 2) and female DNA (lanes 3 and 4); GM06318B (active X chromosome-containing hybrid cell line) DNA (lane 5); HY70C4T3 and THX88 DNA (lanes 6 and 7); GM06317 and HY853 (Y chromosome-containing hybrid cell lines) DNA (lanes 8 and 9); or GM1416 (48,XXXX human lymphoblastoid cell line) DNA (lane 10). Digestions were performed with EcoRI and the methylation-sensitive enzyme BssHII.