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Use of In Vivo and In Vitro Systems to Select Leishmania amazonensis Expressing Green Fluorescent Protein

Korean Journal of Parasitology 2011년 49권 4호 p.357 ~ 364
 ( Costa Solange dos Santos ) - Brazil Universidade Estadual de Campinas Caixa Postal 6109 Department of Animal Biology

 ( Golim Marjorie de Assis ) - Brazil Universidade Estadual de Campinas Evolution and Bioagents, Biology Institute Department of Genetics
 ( Rossi-Bergmann Bartira ) - Universidade Estadual Paulista School of Medicine Faculty of Medicine Blood Center
 ( Costa Fabio Trindade Maranhao ) - Brazil Universidade Federal do Rio de Janeiro Carlos Chagas Filho Institute
 ( Giorgio Selma ) - Brazil Universidade Estadual de Campinas Caixa Postal 6109 Department of Animal Biology

Abstract


Various Leishmania species were engineered with green fluorescent protein (GFP) using episomal vectors that encoded an antibiotic resistance gene, such as aminoglycoside geneticin sulphate (G418). Most reports of GFP-Leishmania have used the flagellated extracellular promastigote, the stage of parasite detected in the midgut of the sandfly vector; fewer studies have been performed with amastigotes, the stage of parasite detected in mammals. In this study, comparisons were made regarding the efficiency for in vitro G418 selection of GFP-Leishmania amazonensis promastigotes and amastigotes and the use of in vivo G418 selection. The GFP-promastigotes retained episomal plasmid for a prolonged period and G418 treatment was necessary and efficient for in vitro selection. In contrast, GFP-amastigotes showed low retention of the episomal plasmid in the absence of G418 selection and low sensitivity to antibiotics in vitro. The use of protocols for G418 selection using infected BALB/c mice also indicated low sensitivity to antibiotics against amastigotes in cutaneous lesions.

키워드

Leishmania amazonensis; green fluorescent protein; macrophage; geneticin
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