Analysis of genomic instability by fluorescence DNA sequencer

: Within the recent four years, there have been substantial advances in understanding the molecular pathogenesis of HNPCC. As one of the result of investigation, microsatellite instability has been observed in hereditary non-polyposis colorectal cancer (HNPCC) and other sporadic cancers. Moreover, there is strong supporting evidence that mismatch repair genes play a role in HNPCC. Here, we present our investigational results and discuss possible molecular mechanisms governing DNA mutation and genomic instability, leading to the development of neoplasm. We investigated replication error (RER) of 4 microsatellite markers (dinucleotide repeats) in 131 patients with colorectal cancer (10 met the criteria of HNPCC group B), using fluorescence-based DNA sequencer. We detected RER positivity (at more than two loci) in 12 of 131 patients (9.2%). PCR-SSCP and direct sequencing of the mismatch repair genes, hMSH2 and hMLH1, revealed that two patients in HNPCC group B had germline mutations of hMSH2. The fluorescence-based techniques, such as the present RER analysis do not require radioactive materials and specialized rooms, and can easily be performed in a clinical laboratory.