PMID: 16637276Apr 28, 2006Paper

Characterization of the universal Russian reagent sets for real-time PCR and its application for molecular oncodiagnostic

Molekuliarnaia biologiia
O Iu ManzeniukA B Poltaraus

Abstract

Universal Russian reagents for real time PCR were tested and compared with reference reagents provided by foreign companies. Testing was carried out on plasmids with cloning fragments (DNA-standards) of cDNA with chimeric (fusion) gene PML-RARalpha. Values of amplification efficiency of Russian and foreign reagents were measured on samples with serial dilutions (30-300000 copies) of cloned cDNA fragments of PML-RARalpha and internal control gene ABL. Amplification efficiencies of Russian and foreign reagents were found to be close one to another. Russian universal reagent kit RealityTM and ABI TaqMan Core Reagent Kit have amplification efficiencies 1.919 and 1.929, and correlation coefficients of copy numbers PML-RARalpha0.999 and 0.996, respectively. These values were determined by construction of a standard curve. To verify these results we studied also the samples of cDNA from blood and bone marrow of patients with acute promyelocytic leukemia. All samples posses translocation t(15;17), and appropriate chimeric gene PML-RARalpha. copy number in 1 microg of total RNA was in range 5.86 x 10(4)-8.315 x 10(5) before chemotherapy. No symptoms of minimal residual disease were found after 3.5 months since chemotherapy - fusion gene...Continue Reading

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