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  • Title: Rapid detection of PML-RARA fusion gene by novel high-speed droplet-reverse transcriptase-polymerase chain reaction: possibility for molecular diagnosis without lagging behind the morphological analyses.
    Author: Sueki A, Matsuda K, Taira C, Yamaguchi A, Koeda H, Takagi F, Kobayashi Y, Sugano M, Honda T.
    Journal: Clin Chim Acta; 2013 Jan 16; 415():276-8. PubMed ID: 23159843.
    Abstract:
    BACKGROUND: Acute promyelocytic leukemia (APL) is an aggressive disease requiring prompt diagnosis and treatment. Rapid detection of the PML-RARA fusion gene provides the molecular basis for a highly effective therapy with all-trans retinoic acid. We developed a rapid assay by novel droplet-reverse transcriptase-polymerase chain reaction (droplet-RT-PCR) for the detection of the PML-RARA fusion gene in APL patients. METHODS: RNA was extracted from 7 samples obtained from 5 APL patients with the PML-RARA fusion gene confirmed by nested RT-PCR and fluorescence in situ hybridization. Using these 7 samples, we evaluated the reaction time and amplification efficiency of the droplet-RT-PCR. RESULTS: Using the droplet-RT-PCR, we could detect the PML-RARA fusion gene in all 7 samples. The reaction time for 50 cycles of droplet-RT-PCR was 27 min. The amplification by the droplet-RT-PCR assay was considered positive for the PML-RARA fusion gene in less than 22 min, at the point when the fluorescence exceeded the threshold level. CONCLUSIONS: Our novel droplet-RT-PCR assay is specific for the detection of the PML-RARA fusion gene and has a markedly reduced reaction time. Thus, the novel droplet-RT-PCR assay contributes to the rapid diagnosis of APL without lagging behind the morphological assessment.
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