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Instituto Oswaldo Cruz, Ministério da Saúde, Memórias do Instituto Oswaldo Cruz., 8(109), p. 1081-1085

DOI: 10.1590/0074-0276140238

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Construction of three new Gateway® expression plasmids for Trypanosoma cruzi

Journal article published in 2014 by Victoria L. Alonso ORCID, Alonso,Victoria L., Carla Ritagliati, Ritagliati,Carla, Pamela Cribb, Cribb,Pamela, Esteban C. Serra, Serra,Esteban C.
This paper is made freely available by the publisher.
This paper is made freely available by the publisher.

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Abstract

We present here three expression plasmids for Trypanosoma cruzi adapted to the Gateway® recombination cloning system. Two of these plasmids were designed to express trypanosomal proteins fused to a double tag for tandem affinity purification (TAPtag). The TAPtag and Gateway® cassette were introduced into an episomal (pTEX) and an integrative (pTREX) plasmid. Both plasmids were assayed by introducing green fluorescent protein (GFP) by recombination and the integrity of the double-tagged protein was determined by western blotting and immunofluorescence microscopy. The third Gateway adapted vector assayed was the inducible pTcINDEX. When tested with GFP, pTcINDEX-GW showed a good response to tetracycline, being less leaky than its precursor (pTcINDEX).