ANALYSIS OF T-CELL RECEPTOR-GAMMA GENE REARRANGEMENTS BY DENATURING GRADIENT GEL-ELECTROPHORESIS OF GC-CLAMPED POLYMERASE CHAIN-REACTION PRODUCTS - CORRELATION WITH TUMOR-SPECIFIC SEQUENCES

We describe a modified denaturing gradient gel electrophoresis (DGGE) procedure with a 40-nucleotide GC clamp in the polymerase chain reacti on to improve resolution in amplifying T cell receptor-gamma (TCR-gamm a) rearrangements. DNA from 46 cases of lymphoblastic leukemia/lymphom a, 5 T cell lines, 2 B cell lines, 7 normal lymphocytes, and 3 cases o f Hodgkin's disease was amplified by polymerase chain reaction. In add ition, 20 cases of paraffin-embedded T cell lymphomas and 5 cases of r eactive hyperplasia were also studied. Clonal TCR-gamma rearrangements were identified on DGGE by the presence of a predominant band. Result s obtained from 5 T cell lines and 12 lymphoblastic leukemia/lymphomas containing known TCR-gamma gene rearrangements revealed 100% concorda nce in detecting clonal rearrangements between DGGE and traditional So uthern blot analysis. Of the remaining 34 lymphoblastic leukemia/lymph oma cases studied by DGGE alone, 30 were positive. DGGE analysis of 10 lymphoblastic leukemia/lymphoma cases with known group IV gamma to J gamma 1 or J gamma 2 rearrangement sequences confirmed that the electr ophoretic migration was dependent on the tumor-specific rearranged TCR -gamma sequence. In addition, 17 of 20 cases of paraffin-embedded T ce ll lymphomas were positive by DGGE, 6 of which had the clonal populati on also identified in fresh tissue DNA. DGGE analysis of GC-clamped po lymerase chain reaction products can provide a way to more accurately detect TCR-gamma clonality of lymphoid tumors and can be applied to ar chival tissues.