| Popis: |
Prognosis of patients with acute myeloid leukemia (AML) is not satisfactory. Long time remission can be achieved only in 25-40% children with AML. The aim of the study is to improve diagnosis of AML in cases characterized by specific chromosomal translocations t(8;21) and t(15;17). These translocations can be efficiently detected using reverse transcriptase-polymerase chain reaction (RT-PCR). To determine optimal conditions for detection of the t(8;21) and t(15;17) translocations in human cells, we performed a detail study of experimental conditions for RT-PCR on human cell lines NB-4 and KASUMI-1. For detection of t(8;21) using Pfx polymerase, the optimal PCR conditions included magnesium ion concentration in the range of 0.6 to 1 mM, 0.4 microM primer concentration, 5 degrees C annealing temperature and 1 microl of template cDNA. For detection of t(15;17) using the Pfx polymerase, the optimal conditions included 1 mM magnesium ion concentration, 0.6 microM primer concentration, 63 degrees C-66 degrees C annealing temperature and 2 microl of template cDNA. The results proved to be valid for clinical diagnostics of these chromosomal aberrations in blood and/or bone marrow samples of AML patients. |
