We have developed an RLGS-based scanning system to detect DNA alteration in tumor tissues, using 575 mapped spots/loci in a single gel. This system is very powerful for screening and identifying not only loss of heterozygosity (LOH) but also DNA methylation change. In this study, we applied this system to search for the LOH of hepatoma from an interspecific F1 hybrid between Mus spretus and C57BL/6 with SV40 early T antigen transgene connected to a mouse major urinary protein enhancer/promoter. Comparing the RLGS profiles of each tumor to that of the normal tissue showed significant LOH in chromosomes 1, 5, 7 and 13.