Biochemical and Biophysical Research Communications
Trypanosoma cruzi: TcRAB7 protein is localized at the Golgi apparatus in epimastigotes
Section snippets
Materials and methods
Parasites. T. cruzi clones, CL Brener, and Dm28c, were used. Epimastigotes were cultivated at 28 °C in liver infusion tryptose (LIT) liquid medium supplemented with 10% fetal calf serum [18]. Stably transformed epimastigote clones, transfected with either control plasmid pTEX [19] or with pTAG, a plasmid that leads to over-expression of a hemagglutinin (HA) epitope-tagged TcRAB7 protein [17], were maintained in the presence of 500 μg/ml of G418 (Geneticin, Gibco, Grand Island, NY).
Nucleic acid
Characterization of an immune serum against TcRAB7 protein of T. cruzi
The immune serum was prepared against the synthetic peptide corresponding to the non-conserved carboxy-terminal 14-amino acid sequence of TcRAB7. Immunoblotting experiments were performed using the recombinant TcRAB7 protein as antigen. Total immune serum recognized a protein with molecular weight corresponding to GST-fused TcRAB7 protein (around 56 kDa, Fig. 1A) on Western blots of the fusion protein from total extract of E. coli. The total immune serum recognized neither bacterial proteins nor
Acknowledgments
We thank Rosane Silva and Turán P. Ürményi for critical reading of the manuscript and César Félix Schmidt, Cláudio Nunes Pereira, and Sebastisião Cruz for technical assistance. This work was supported by grants from the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq), Fundação Coordenação de Aperfeiçoamento de Pessoal de Nı́vel Superior (CAPES), Financiadora de Estudos e Projetos (FINEP), Fundação José Bonifácio (FUJB), Fundação Carlos Chagas Filho de Apoio à Pesquisa no
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