SELECTION OF PROBES FOR FLUORESCENCE IN-SITU HYBRIDIZATION ANALYSIS BY DIFFERENTIAL DISPLAY POLYMERASE CHAIN-REACTION OF MESSENGER-RNA FROMRHABDOMYOSARCOMA

Rhabdomyosarcoma (RMS) is a malignancy of skeletal muscle derivation e ncompassing two major subtypes, embryonal and alveolar, which differ i n clinical behavior and genetic markers. Because RMS is a relatively c ircumscribed tumor system for which the beginnings of a molecular gene tic framework are in place, it becomes an ideal model for the applicat ion of improved methods of molecular genetic analysis. We have applied the technique known as differential display polymerase chain reaction (DD-PCR) to characterize expression of RNA in rhabdomyosarcoma subtyp es. Our studies have shown that DD-PCR generates a characteristic elec trophoretic profile that can be used to isolate subtype specific probe s fluorescence in situ hybridization (FISH) analysis. We have isolated two cDNA fragments and obtained clones suitable for FISH mapping to m etaphase chromosomes. One probe was mapped to the centromeric region o f human chromosome 22 and the other probe to the human chromosome band 6q25-26. This approach demonstrates the utility of DD-PCR as a techni que for isolating novel cDNA expressed in tumors and their subsequent use as probes for FISH analysis. As more genes ore identified by DD-PC R and their roles in tumorigenesis become defined, they are likely to provide novel targets for future molecular cytogenetic analysis. (C) E lsevier Science Inc., 1996