Journal
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
Volume 408, Issue 4, Pages 1079-1094Publisher
SPRINGER HEIDELBERG
DOI: 10.1007/s00216-015-9204-2
Keywords
BCR-ABL; Molecular diagnostics; Philadelphia chromosome; Cancer diagnostics; Digital PCR; Chronic myeloid leukemia
Funding
- Natural Sciences and Engineering Research Council of Canada (NSERC)
- Canadian Institutes of Health Research (CIHR)
- Genome BC
- Canada Research Chair
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Formed from a reciprocal translocation t(9:22)(q34;q11) of genetic material between the long arms of human chromosomes 9 and 22, the constitutively active breakpoint cluster region (BCR) Abelson 1 (ABL) tyrosine kinase BCR-ABL is known to be causative of chronic myelogenous leukemia (CML). In 98 % of CML patients harboring the t(9:22)(q34;q11) translocation, known as the Philadelphia chromosome, the chimeric BCR-ABL oncogene is created through cleavage of the BCR gene within its major breakpoint region (M-BCR) and breakage of the ABL gene within a 100-kbp region downstream of exon 2a. Clinical detection of the fused BCR-ABL oncogene currently relies on direct visualization by fluorescence in situ hybridization (FISH), a relatively tedious assay that typically offers a detection limit of ca. 2 %. Here, we describe a novel assay that uses droplet digital PCR (ddPCR) technology to reliably measure M-BCR status and the presence of BCR-ABL. When applied to cell-line models of CML, the assay accurately quantifies BCR-ABL frequency to a detection limit of 0.25 %. It therefore offers improved specificity relative to FISH, and may allow identification of variant translocation patterns, including derivative chromosome 9 deletions.
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