Rapid amplification of immunoglobulin heavy chain switch (IGHS) translocation breakpoints using long-distance inverse PCR.
Molecular cloning of immunoglobulin heavy chain (IGH) translocation breakpoints identifies genes of biological importance in the development of normal and malignant B cells. Long-distance inverse PCR (LDI-PCR) was first applied to amplification of IGH gene translocations targeted to the joining (IGHJ) regions. We report here successful amplification of the breakpoint of IGH translocations targeted to switch (IGHS) regions by LDI-PCR. To detect IGHS translocations, Southern blot assays using 5' and 3' switch probes were performed. Illegitimate Smu rearrangements were amplified from the 5' end (5'Smu LDI-PCR) from the alternative derivative chromosome, and those of Sgamma or Salpha were amplified from the 3' end (3'Sgamma or 3'alpha LDI-PCR) from the derivative chromosome 14. Using a combination of these methods, we have succeeded in amplifying IGHS translocation breakpoints involving FGFR3/MMSET on 4p16, BCL6 on 3q27, MYC on 8q24, IRTA1 on 1q21 and PAX5 on 9p13 as well as BCL11A on 2p13 and CCND3 on 6p21. The combination of LDI-PCR for IGHJ and IGHS allows rapid molecular cloning of almost all IGH gene translocation breakpoints.
Sonoki
T
T
Academic Haematology and Cytogenetics, Institute of Cancer Research, UK.
Willis
T G
TG
Oscier
D G
DG
Karran
E L
EL
Siebert
R
R
Dyer
M J S
MJ
D001483:Base Sequence; D015139:Blotting, Southern; D002352:Carrier Proteins / Q000235:genetics; D019457:Chromosome Breakage / Q000235:genetics*; D002883:Chromosomes, Human, Pair 14; D003001:Cloning, Molecular; D004268:DNA-Binding Proteins / Q000235:genetics; D015321:Gene Rearrangement; D005812:Genes, Switch / Q000235:genetics*; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007134:Immunoglobulin Switch Region / Q000235:genetics*; D007148:Immunoglobulin mu-Chains / Q000235:genetics; D008223:Lymphoma / Q000235:genetics*; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000235:genetics; D051757:PAX5 Transcription Factor; D016133:Polymerase Chain Reaction; D011505:Protein-Tyrosine Kinases / Q000235:genetics; D011518:Proto-Oncogene Proteins / Q000235:genetics; D051560:Proto-Oncogene Proteins c-bcl-6; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics; D051498:Receptor, Fibroblast Growth Factor, Type 3; D017468:Receptors, Fibroblast Growth Factor / Q000235:genetics; D012097:Repressor Proteins; D014157:Transcription Factors / Q000235:genetics; D014178:Translocation, Genetic; D014407:Tumor Cells, Cultured
C486563:BCL11A protein, human; D002352:Carrier Proteins; D004268:DNA-Binding Proteins; D007143:Immunoglobulin Heavy Chains; D007148:Immunoglobulin mu-Chains; D009687:Nuclear Proteins; D051757:PAX5 Transcription Factor; C497281:PAX5 protein, human; D011518:Proto-Oncogene Proteins; D051560:Proto-Oncogene Proteins c-bcl-6; D016271:Proto-Oncogene Proteins c-myc; D017468:Receptors, Fibroblast Growth Factor; D012097:Repressor Proteins; D014157:Transcription Factors; C496365:FGFR3 protein, human; D011505:Protein-Tyrosine Kinases; D051498:Receptor, Fibroblast Growth Factor, Type 3