RESUMO
The use of accurate and reliable techniques for sex determination of wild birds is of special importance in captive breeding programs, especially in birds with monogamous, aggressive behavior, with absence of copulation, and with a low hatching rate. Using PCR, we evaluated the relative efficacy of primers HPF/HPR and CHD1Wr/NP/CHD1Zr in the amplification of the chromo-helicase-DNA binding 1 (CHD1) gene for sex determination in Penelope albipennis and 8 other species of cracids, 4 species of falconids, 4 species of accipitrids, and 3 species of psittacines. Primer effectiveness was compared using previously sexed bird samples. The HPF/HPR primer set was found to demonstrate a better performance and reliability. Therefore, these primers should be used to determine the sex of juvenile birds to avoid or minimize incompatibilities during the selection of potential breeding pairs.
Assuntos
Animais Selvagens/genética , Aves/genética , Análise para Determinação do Sexo , Animais , DNA/isolamento & purificação , Primers do DNA/metabolismo , Feminino , Genoma , Masculino , Peru , Reação em Cadeia da Polimerase , Especificidade da EspécieRESUMO
El presente trabajo informa de la purificación y caracterización bioquímica y biológica de la fosfolipasa A2 (PLA2) de Lachesis muta (Linnaeus, 1766). La purificación se realizó por cromatografía liquida (CL) usando CM-Sephadex C-50 y Sephadex G-50, obteniéndola al estado homogéneo con un peso molecular de 18749 Da. Los ensayos con PLA2 realizados sobre fosfolípidos de yema de huevo y lecitina comercial, mostraron que los agentes EDTA, PMSF, glutatión y cisteína, inhibieron la actividad con valores mayores al 50%. La PLA2 de L. muta produjo un notable efecto anticoagulante, observándose un retardo de 2'30' en el tiempo de coagulación con 9,6 microgramos de la enzima. La hemólisis indirecta sobre eritrocitos humanos dio un equivalente de 4,35 microgramos como dosis hemolítica media (HD50). Los valores de dosis edemática media y dosis miotóxica mínima fueron de 91,5 microgramos y 125,89 microgramos/mL respectivamente; valores por debajo de PLA2 de otros venenos. No se registró actividad hemorrágica directa. Las pruebas de inmunodifusión e inmunoelectroforésis revelaron que PLA2 de L. muta tuvo reactividad inmunogénica contra el antiveneno lachésico monovalente (INS-Perú). Sin embargo, la neutralización por el antiveneno fue parcial.
In the present study, phospholipase A2 (PLA2) from Lachesis muta (Linnaeus, 1766), is isolated, purified and characterized biochemically and biologically. Purification was performed by liquid chromatography (LC) using CM-Sephadex C-50 and Sephadex G-50, homogenized enzyme had a molecular weight of 18749 Da. Trials with egg yolk phospholipids, and commercial lecithin showed that EDTA, PMSF, glutathione and cysteine inhibited the activity with values greater than 50%. The PLA2 had a significant anticoagulant effect, showing a delay of 2'30" on the coagulation time with 9.6 microgramos of the enzyme. The indirect impact on human erythrocyte hemolysis gave an equivalent of 4.35 microgramos as HD50. Mean edematic dose and minimum myotoxic dose were 91.5 mg and 125.89 mg / mL respectively, these values were below enzymes phospholipase A2 from others poisons. There was no hemorrhagic activity. Immunodiffusion tests and immunoelectrophoresis revealed that the PLA2 of L. muta was immunogenic reactivity against lachesic monovalent antivenom (INS-Peru). However, the neutralization by the antivenom was partial.
