title: [PCR-RFLP/Hsp70 for identification and tipification of leishmania from the tropical region].
abstract: The optimization of the PCR conditions for amplification of the gene coding for the 70 kDa (HSp70) heat shock protein as well as the analysis of the restriction fragment length polymorphism (RFLP) were carried out. DNA from a reference strain of Leishmania mexicana was used as template. Analytical sensitivity and specificity, and reproducibility of PCR using DNA from L. mexicana, Lamazonensis, L. guyanensis and L. lainsoni were determined. A 1.3 kp band was obtained, which confirmed gene amplification. The band patterns derived from Haelll enzyme digestion allowed differentiating several species. L. guyanensis and L. lainsoni were different from each other, while L. mexicana and L. amazonensis, which shared a common pattern, were different from the other two species. Analytical sensitivity and specificity were adequate. The enzymatic restriction of the PCR product made it possible to differentiate Leishmania spp. from T. cruzi. The feasibility of identifying and typifying species from the American continent through PCR-RFLP/Hsp70 and of using enzymatic restriction of amplified product to distinguish Leishmania spp. from Trypanosornma cruzi was shown. This was the first step in implementing these molecular methods in the reference laboratory of the Institute.
authors: Instituto de Medicina Tropical Pedro Kourí. amontalvo@ipk.sld.cu
authors: Ana
authors: A
authors: Margarita Montalvo
authors: Jorge
authors: J
authors: Fraga
authors: Jaqueline
authors: J
authors: Aylema Romero
authors: Lianet
authors: L
authors: Monzote
authors: Ing
authors: I
authors: Ivon Montano
authors: Jean Claude
authors: JC
authors: Dujardin
mesh_terms: D015373:Bacterial Typing Techniques / Q000379:methods / Q000592:standards*; D016054:DNA, Protozoan / Q000032:analysis / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics*; D007843:Latin America; D007891:Leishmania / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction / Q000379:methods / Q000592:standards*; D012150:Polymorphism, Restriction Fragment Length; D015800:Protozoan Proteins / Q000235:genetics*; D011550:Pseudomonas aeruginosa / Q000235:genetics; D012015:Reference Standards; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D013045:Species Specificity; D014122:Toxoplasma / Q000235:genetics; D014349:Trypanosoma cruzi / Q000235:genetics
chemical_list: D016054:DNA, Protozoan; D018840:HSP70 Heat-Shock Proteins; D015800:Protozoan Proteins; D015252:Deoxyribonucleases, Type II Site-Specific; C024403:GGCC-specific type II deoxyribonucleases