Easy detection of all T cell receptor gamma (TCRG) gene rearrangements by Southern blot analysis: recommendations for optimal results

Abstract

Southern blot analysis of T cell receptor (TCR) gene rearrangements has proven to be a helpful tool to establish clonality in T cell leukemias and lymphomas. To improve the detection of clonal TCR gamma (TCRG) gene rearrangements by Southern blot analysis, we designed four new Jγ probes and determined the most optimal restriction enzymes to be used with these probes. Based on detailed analysis of the sequences as well as on hybridization experiments with the TCRGJ21 probe, the Jγ1.2 and Jγ2.1 downstream areas were found to be highly homologous, suggesting that during evolution the duplication of the Jγ region was followed by deletion of the tentative Jγ2.2 gene segment. Southern blot analysis of 51 T cell acute lymphoblastic leukemias (T-ALL) revealed that all TCRG gene rearrangements can be detected by use of the TCRGJ13 probe in EcoRI digests and the TCRGJ21 probe in PstI digests. Additional probes and digests allow a more precise identification of the exact type of TCRG gene rearrangements in the majority of cases. Almost 90% of the TCRG gene rearrangements in T-ALL involved the Jγ2 region (16% Jγ2.1 and 72% Jγ2.3), whereas Jγ1 region rearrangements were particularly found in TCRγδ⁺ T-ALL. This information has implications for design of primer sets for PCR analysis at diagnosis and for PCR target choice in detection of minimal residual disease during follow-up of T-ALL patients.