High-resolution detection of loss of heterozygosity of dinucleotide microsatellite markers.

Dinucleotide microsatellite markers are frequently investigated to study inheritance, genetic stability, and allele frequency distribution in a wide variety of genetic disorders. Previous studies have encountered significant problems regarding resolution and detection of dinucleotide, microsatellites. In this study, a useful method to investigate loss of heterozygosity (LOH) of dinucleotide microsatellite markers is described that involves the use of nondenaturing (Spreadex) submerged gel electrophoresis and SYBR Green I nucleic acid staining. This method omits the gel casting step and the use of hazardous radioactive materials frequently used in many microsatellite studies that employ polyacrylamide gel nucleic acid denaturation analysis. Using this method, 62 patients' paired tumor and normal samples were investigated to detect allele deletions in a region of chromosome 7q31.1, which is believed to harbor a tumor suppressor gene. Interpretable results were obtained in all cases. These results were compared to those attained using ABI Prism Genetic Analyzer 310 and Gene-Scan. There were no discrepancies in results obtained between the two assays. The Spreadex system is cheap, does not require larger equipment costs, and may prove to be a useful system for high-throughput investigation of microsatellites. It may have diagnostic significance and also prove useful if applied to population-based genomic screening and linkage analysis.

Transcriptional gene expression profiles of oesophageal adenocarcinoma and normal oesophageal tissues.

The molecular events associated with the development of adenocarcinoma of the oesophagus are not well understood. Gene expression associated with oesophageal adenocarcinoma was investigated using a cDNA array containing 1,176 human cancer-associated genes. Approximately 59% of the genes were expressed at detectable levels with 15 genes (1.3%) exhibiting differential (> 2.5-fold) expression in either normal oesophagus or adenocarcinoma tissue. Nine genes were up-regulated in oesophageal adenocarcinoma tissue (matrix metalloproteinase 11 (MMP11), ornithine decarboxylase (ODC), cytokeratins 8 and 18, integrin alpha 3 (ITGA3), integrin alpha 6 (ITGA6), BIGH3 (transforming growth factor beta-induced), beta-catenin and CDC25B (M-phase inducer phosphatase 2)). Six genes were down-regulated in adenocarcinoma tissue (cytokeratin 4, plasminogen activator inhibitor 2 (PAI-2), interleukin 1 receptor antagonist (IRAP), cytokeratin 13/15/17, MAD and retinoic acid receptor gamma 1 (RARG)). Many of these differentially expressed genes influence cell-cell adhesion, cell-extracellular matrix and composition, transcriptional activation and cell cycle progression and are likely to contribute to development of oesophageal adenocarcinoma.