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|Title:||Transient transfection and expression of foreign and endogenous genes in the intracellular stages of Trypanosoma cruzi|
|Authors:||Padmanabhan, Prasad K.|
Polidoro, Rafael B.
Barteneva, Natasha S.
Gazzinelli, Ricardo Tostes
Burleigh, Barbara A.
|Affilliation:||Harvard School of Public Health. Department of Immunology and Infectious Diseases. Boston, MA, United States|
Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia e Departamento de Parasitologia. Belo Horizonte, MG, Brasil
Harvard Medical School. Boston Children's Hospital and Department of Pediatrics. Cellular and Molecular Medicine Program. Boston, MA, United States
Universidade Federal de Minas Gerais. Instituto de Ciências Biológicas. Departamento de Bioquímica e Imunologia e Departamento de Parasitologia. Belo Horizonte, MG, Brasil/ Fundação Oswaldo Cruz. Centro de Pesquisas René Rachou. Belo Horizonte, MG, Brazil/University of Massachusetts Medical School. Division of Infectious Disease and Immunology. MA, United States
Harvard School of Public Health. Department of Immunology and Infectious Diseases. Boston, MA, United States
|Abstract:||The capacity for rapid localization of epitope-tagged or fluorescent fusion proteins in cells is an important tool for biological discovery and functional analysis. For Trypanosoma cruzi, the protozoan parasite that causes human Chagas disease, visualization of ectopically-expressed proteins in the clinically-relevant mammalian stages is hindered by the necessity to first perform transfection and lengthy selection procedures in the insect vector form of the parasite. Here, we demonstrate the ability to by-pass the insect stage with the delivery of plasmid DNA to non-dividing, tissue culture trypomastigotes such that upon host cell infection, transgenes are expressed and rapidly localized in intracellular T. cruzi amastigotes. The inclusion of a sorting step prior to host cell infection by trypomastigotes greatly enriches (>90%) the number of transgene-expressing amastigotes observed in mammalian host cells. This is a significant methodological advance that has the potential to accelerate the pace of discovery in the Chagas disease field.|
Fluorescence-activated cell sorting
Mammalian cell infection
|Citation:||PADMANABHAN, Prasad K. et al. Transient transfection and expression of foreign and endogenous genes in the intracellular stages of Trypanosoma cruzi. Molecular and Biochemical Parasitology, Vol. 198, n. 2, p. 100–103, 2014|
|Appears in Collections:||MG - IRR - Artigos de Periódicos|
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