Analysis of GGAA-Microsatellites in Ewing Sarcoma

OBJECTIVES:

• To describe the spectrum of GGAA-microsatellite polymorphisms at specific loci in genomic DNA prepared from Ewing sarcoma tumor specimens.
• To determine if there are differences in GGAA-microsatellite polymorphisms in genomic DNA prepared from Ewing sarcoma tumor specimens as compared to non-afflicted European and African normal genomic DNA.
• To determine if GGAA-microsatellite polymorphisms at specific loci in genomic DNA prepared from Ewing's sarcoma tumor specimens correlates with disease outcome in patients treated on COG protocol AEWS0031.
• To determine whether whole-genome amplification introduces alterations in GGAA-microsatellites as compared to non-amplified genomic DNA.

OUTLINE: Genomic PCR is used to amplify the microsatellites in the NR0B1 and GSTM4 promoters. In addition to determining microsatellite size, each microsatellite is sequenced following cloning into the pCR4 vector (Invitrogen) using standard topoisomerase cloning protocols.