Top: Methylation status of chromosome 1 heterochromatin DNA in HCC. Hypomethylation status of satellite 2 (Sat2) DNA of chromosome 1 was studied by Southern blot. DNA extracted from HCC was digested by methyl-sensitive enzyme BstB1 and probed against Sat2. Liver (L) and sperm (Sp) DNAs were used as the methylated (negative control) and hypomethylated (positive control) standards, respectively. The degree of hypomethylation in each lane was quantitated by comparing the ratio intensities of hybridized fragments smaller than 4 kb to those larger than 4-kb molecular weight. DNA samples with ratios <0.7, 0.7 to 1.1, and >1.1 were considered to have a normal level of methylation, moderate level of Sat2 hypomethylation, and extensive hypomethylation, respectively. Bottom: Molecular cytogenetic analysis on HCC. A: SKY analysis on H29 revealed copies of chromosome 1 with moderate to extensive decondensation of the heterochromatic region. Repetitive DNA sequences within the heterochromatic region had been suppressed by Cot-1 DNA but the reverse DAPI image indicated fragility of the segment. B: CGH analysis on HCC revealed frequent 1q copy number gain. Visual inspection of hybridized chromosome often indicated a strong staining green region of tumor DNA on the q-arm and a suggested breakpoint within 1q12. The figure shown illustrates the 1q copy gain detected in case H2. Fluorescent ratio intensities is plotted alongside the chromosome ideogram and the ratio profile of 16 chromosomes (n = 16, pink line) is depicted with 95% confidence interval (gold lines). C: SKY analysis on H29 supported CGH finding for the presence of balanced chromosome 1. The finding of chromosome 1p and 1q translocation, t(1;17)(p10;p10) and t(1;3)(q10;p10), suggested no net gain or loss of chromosome 1 material. D: Chromosome 1 translocation in case H34. Classified image is displayed alongside the reverse DAPI image. SKY analysis indicated a translocation between chromosomes 1 and 5, t(1;5)(p22;?q), with a p-arm breakpoint on band 1p22. E: Classified karyotype of chromosome 1 in case H32. Reverse DAPI-banding pattern indicated a translocation between chromosomes 1 and 8, t(1;8)(p13;q11.2), with a chromosome 1 breakpoint on p13.