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Memórias do Instituto Oswaldo Cruz
Fundação Oswaldo Cruz, Fiocruz
ISSN: 1678-8060
EISSN: 1678-8060
Vol. 95, No. 1, 2000, pp. 111-114
Bioline Code: oc00018
Full paper language: English
Document type: Research Article
Document available free of charge

Memórias do Instituto Oswaldo Cruz, Vol. 95, No. 1, 2000, pp. 111-114

 en Short Communication - Colony Polymerase Chain Reaction of Stably Transfected Trypanosoma cruzi Grown on Solid Medium
Wagner G dos Santos; Ivelina Metcheva & Gregory A Buck

Abstract

Tools for the genetic manipulation of Trypanosoma cruzi are largely unavailable, although several vectors for transfection of epimastigotes and expression of foreign or recombinant genes have been developed. We have previously constructed several plasmid vectors in which recombinant genes are expressed in T. cruzi using the rRNA promoter. In this report, we demonstrate that one of these vectors can simultaneously mediate expression of neomycin phosphotransferase and green fluorescent protein when used to stably transfect cultured epimastigotes. These stably transfected epimastigotes can be selected and cloned as unique colonies on solid medium. We describe a simple colony PCR approach to the screening of these T. cruzi colonies for relevant genes. Thus, the methodologies outlined herein provide important new tools for the genetic dissection of this important parasite.

Keywords
Trypanosoma cruzi - stable transfection - Green Fluorescent Protein - neomycin phosphotransferase

 
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