Refinement of the BCL2/immunoglobulin heavy chain fusion gene in t(14;18)(q32;q21) by polymerase chain reaction amplification for long targets

The t(14;18)(q32;q21) translocation, involving the BCL2 gene and junctional segments (JH) of the immunoglobulin heavy chain gene (IGH), constitutes the most common chromosomal translocation in non‐Hodgkin's lymphoma of B‐cell type. Although the breakpoints in BCL2 are largely clustered within the major breakpoint region (MBR) and minor cluster region (mcr), it is known that some breakpoints map away from these regions, resulting in negative amplification of the junctional sequence by polymerase chain reaction (PCR) for 20 kb targets, which covered the intervening region between the MBR and mcr. Restriction analysis of the LD‐PCR products revealed that breakpoints in 33 cases fell within the 150 bp‐MBR region, and in 3 cases were within the mcr determined previously by others. In contrast, the breakpoints of the remaining 16 cases were distributed over a large region from the MBR through mcr. Nucleotide sequence analysis of a potential cluster region revealed the presence of an Alu repeat sequence. Restriction analysis of LD‐PCR products with BstEII demonstrated a predominant usage of the JH6 segment (71%) at the BCL2/JH junctions. LD‐PCR using primers for the constant region genes showed that class switch recombination occurred in more than 80% of the IGH genes on the der(14) chromosome. Our study showed that LD‐PCR was capable of detecting virtually any t(14;18) that occurred within the approximately 30 kb region downstream of the MBR, and thus is suitable for initial diagnosis of lymphoma tissues. Furthermore, as amplified fragments obtained by the LD‐PCR contained distinctive regions of BCL2 and IGH, restriction analysis and nucleotide sequencing of the products refined the characteristics of t(14;1