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Memórias do Instituto Oswaldo Cruz
Fundação Oswaldo Cruz, Fiocruz
ISSN: 1678-8060
EISSN: 1678-8060
Vol. 109, No. 8, 2014, pp. 1081-1085
Bioline Code: oc14163
Full paper language: English
Document type: Note
Document available free of charge

Memórias do Instituto Oswaldo Cruz, Vol. 109, No. 8, 2014, pp. 1081-1085

 en Construction of three new Gateway® expression plasmids for Trypanosoma cruzi
Alonso, Victoria L.; Ritagliati, Carla; Cribb, Pamela & Serra, Esteban C.

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).

Keywords
Trypanosoma cruzi; expression vectors; Gateway® system; tandem affinity purification; pTcINDEX

 
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