Methylation of the TLX1 gene in ductal carcinoma in situ and invasive primary breast tumors. Six cases of ductal carcinoma in situ (DCIS), 16 cases of stage I tumors, and eight cases of stage II tumors were analyzed. After sodium bisulfite treatment, the CpG island within the TLX1 gene was amplified with suitable primers and subjected to the BstUI combined bisulfite restriction analysis assay. Digested fragments on the gel are indicative of methylated BstUI restriction sites (5'-CGCG) within the CpG island. Vertical arrows indicate tumor-specific methylation in the target sequence. N/T, normal/tumor pairs; M, DNA marker; Meth. DNA, HeLa DNA methylated in vitro with SssI methyltransferase, serving as a positive control.