Avaliação da estabilidade do plasmídeo pQE-30 e expressão do antígeno 503 de Leishmania i. chagasi em Escherichia coli M15 em diferentes concentrações de IPTG e temperaturas de cultivo
Visceral leishmaniasis is an infectious-parasitic disease caused by protozoa of the genus Leishmania, which can be lethal when there is no adequate treatment. It is estimated that there are 12 million people infected by the disease, but unfortunately there is no vaccine capable of preventing the dis...
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| Formato: | Dissertação |
| Idioma: | por |
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| Endereço do item: | https://repositorio.ufrn.br/jspui/handle/123456789/25113 |
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| Resumo: | Visceral leishmaniasis is an infectious-parasitic disease caused by protozoa of the genus Leishmania, which can be lethal when there is no adequate treatment. It is estimated that there are 12 million people infected by the disease, but unfortunately there is no vaccine capable of preventing the disease. In addition, the treatment of the disease presents high cost and high toxicity. Therefore, we must highlight the research of purified antigenic components that can be used as a tool to obtain vaccines or tests for specific diagnosis. In this context, the present work aims to evaluate the influence of the concentration of IPTG during the induction and the influence of culture temperature on 503 antigen expression and on the stability of plasmid pQE-30 in Escherichia coli. All assays were performed in a rotary incubator and made in duplicate. First, IPTG induced cultures were carried out at concentrations of: 0.01 mM; 0.1 mM; 0.5 mM; 1.0 mM and 1.5 mM at 37 °C. The concentration of the inducer did not affect the stability of the plasmid and when induced with 1.0 mM, the 503 antigen expression was maximal 0.101 ± 0.004 g / L. It was possible to verify that the lowest concentration of IPTG used was enough to achieve expression of the recombinant protein. Thereafter, the concentration of IPTG was fixed at 1.0 mM and the cultivation temperatures of 27, 32 and 42 °C were evaluated. These assays were compared with the previous assay performed at 37 °C. Temperature influenced the stability of the plasmid, and the lower the culture temperature, the greater the stability. According to the results, the temperature of 37 °C obtained the highest expression of 503 antigen, being considered therefore, the optimal conditions for 503 antigen production in a rotary incubator. |
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