TY - JOUR
T1 - Novel 5' exonuclease-based real-time PCR assay for the detection of t(14;18)(q32;q21) in patients with follicular lymphoma
AU - Luthra, Rajyalakshmi
AU - McBride, J. Adelia
AU - Cabanillas, Fernando
AU - Sarris, Andreas
PY - 1998/7
Y1 - 1998/7
N2 - The exonuclease-based real-time polymerase chain reaction (PCR) exploits 5'→3' exonuclease activity of Taq polymerase and measures PCR product accumulation as the reaction proceeds through a dual-labeled fluorogenic probe. The utility of this exonuclease-based PCR assay as a rapid alternative to conventional PCR for follicular lymphoma-associated t(14;18)(q32; q21) was evaluated in this study. The specificity of the assay for t(14;18) involving bcl-2 and immunoglobulin heavy-chain joining region (JH) genes was assessed by analyzing DNA from 53 patients (38 B-cell non-Hodgkin's lymphomas and 15 nonneoplastic proliferations) and correlating the exonuclease PCR data with conventional PCR results. bcl-2/JH fusion sequences were detected by exonuclease-based PCR in 24 of 25 cases shown to be bcl-2 rearranged by conventional PCR. Fusion sequences were not detected in patients who were negative by conventional PCR. The overall concordance between the two assays was 98% (52 of 53 cases concordant positive or negative). In a serial dilution study using t(14;18)-positive cell line DNA, exonuclease-based PCR detected fusion sequences at DNA concentrations of 5 pg, equivalent to 0.6 to 0.8 genomes per reaction. Thus, this study demonstrated that exonuclease- based PCR for t(14; 18) is both specific and highly sensitive. The elimination of the post-PCR amplicon detection steps and the ability to quantitate the input target DNA sequences make this assay ideal for routine diagnostics and monitoring minimal residual disease.
AB - The exonuclease-based real-time polymerase chain reaction (PCR) exploits 5'→3' exonuclease activity of Taq polymerase and measures PCR product accumulation as the reaction proceeds through a dual-labeled fluorogenic probe. The utility of this exonuclease-based PCR assay as a rapid alternative to conventional PCR for follicular lymphoma-associated t(14;18)(q32; q21) was evaluated in this study. The specificity of the assay for t(14;18) involving bcl-2 and immunoglobulin heavy-chain joining region (JH) genes was assessed by analyzing DNA from 53 patients (38 B-cell non-Hodgkin's lymphomas and 15 nonneoplastic proliferations) and correlating the exonuclease PCR data with conventional PCR results. bcl-2/JH fusion sequences were detected by exonuclease-based PCR in 24 of 25 cases shown to be bcl-2 rearranged by conventional PCR. Fusion sequences were not detected in patients who were negative by conventional PCR. The overall concordance between the two assays was 98% (52 of 53 cases concordant positive or negative). In a serial dilution study using t(14;18)-positive cell line DNA, exonuclease-based PCR detected fusion sequences at DNA concentrations of 5 pg, equivalent to 0.6 to 0.8 genomes per reaction. Thus, this study demonstrated that exonuclease- based PCR for t(14; 18) is both specific and highly sensitive. The elimination of the post-PCR amplicon detection steps and the ability to quantitate the input target DNA sequences make this assay ideal for routine diagnostics and monitoring minimal residual disease.
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U2 - 10.1016/S0002-9440(10)65546-0
DO - 10.1016/S0002-9440(10)65546-0
M3 - Article
C2 - 9665466
AN - SCOPUS:0031819880
SN - 0002-9440
VL - 153
SP - 63
EP - 68
JO - American Journal of Pathology
JF - American Journal of Pathology
IS - 1
ER -