Quantification of bcl-2/JH fusion sequences and a control gene by multiplex real-time PCR coupled with automated amplicon sizing by capillary electrophoresis

Beatriz Sanchez-Vega, Francisco Vega, L. Jeffrey Medeiros, Ming S. Lee, Rajyalakshmi Luthra

Research output: Contribution to journalArticle

16 Citations (Scopus)

Abstract

Follicular lymphoma is characterized by the presence of the t(14;18)(q32;q21) chromosomal translocation which juxtaposes the bcl-2 gene at 18q21 with the immunoglobulin heavy chain locus at 14q32. Quantification of t(14;18) carrying cells in FL patients can be achieved by real-time PCR, a highly sensitive technique for evaluating treatment efficacy and minimal residual disease. Despite the many advantages of real-time technology for this purpose, one disadvantage is that current real-time t(14;18) PCR assays amplify a control gene as a normalizer in a separate reaction. Since each PCR reaction has its own kinetics, separate PCR assays for target and control sequences can potentially result in inaccurate quantification of t(14;18)-positive cells. In addition, the real-time t(14;18) PCR assays do not determine the size of the amplified fusion sequence, which is helpful for excluding contamination and is commonly used to demonstrate clonal identity between pre- and post-treatment specimens from a patient. To address these limitations, we designed a multiplex real-time PCR protocol that allows amplification of control and target genes in the same reaction and precise size determination of bcl-2/JH fusion sequences by capillary electrophoresis. This multiplex PCR assay is equally sensitive to previous assays, allows more accurate quantification of bcl-2/JH fusion sequences, and is more convenient.

Original languageEnglish
Pages (from-to)223-229
Number of pages7
JournalJournal of Molecular Diagnostics
Volume4
Issue number4
StatePublished - Nov 1 2002
Externally publishedYes

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Multiplex Polymerase Chain Reaction
Capillary Electrophoresis
Real-Time Polymerase Chain Reaction
Polymerase Chain Reaction
Genes
bcl-2 Genes
Immunoglobulin Heavy Chains
Genetic Translocation
Follicular Lymphoma
Residual Neoplasm
Technology
Therapeutics

ASJC Scopus subject areas

  • Molecular Biology

Cite this

Quantification of bcl-2/JH fusion sequences and a control gene by multiplex real-time PCR coupled with automated amplicon sizing by capillary electrophoresis. / Sanchez-Vega, Beatriz; Vega, Francisco; Medeiros, L. Jeffrey; Lee, Ming S.; Luthra, Rajyalakshmi.

In: Journal of Molecular Diagnostics, Vol. 4, No. 4, 01.11.2002, p. 223-229.

Research output: Contribution to journalArticle

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