Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols

To improve transfection efficiency in Trypanosoma cruzi, we developed a new electroporation protocol and expression vectors which use luciferase and green and red fluorescent proteins as reporter genes. In transient transfections, the electroporation conditions reported here resulted in luciferase e...

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Publicado:	2004
Materias:	alpha tubulin beta tubulin DNA fragment immunoglobulin luciferase red fluorescent protein ribosome protein 5' untranslated region article electroporation epimastigote expression vector gene expression microbial genetics nonhuman priority journal reporter gene Trypanosoma cruzi 5' Untranslated Regions Animals Base Sequence Electroporation Genes, Reporter Genetic Vectors Green Fluorescent Proteins Luciferases Luminescent Proteins Molecular Sequence Data Protozoan Proteins Transfection Tubulin Protozoa Trypanosoma
Acceso en línea:	https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_09320113_v92_n2_p113_DaRocha http://hdl.handle.net/20.500.12110/paper_09320113_v92_n2_p113_DaRocha
Aporte de:	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA) de Universidad de Buenos Aires

id	paper:paper_09320113_v92_n2_p113_DaRocha
record_format	dspace
spelling	paper:paper_09320113_v92_n2_p113_DaRocha2023-06-08T15:53:02Z Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols alpha tubulin beta tubulin DNA fragment immunoglobulin luciferase red fluorescent protein ribosome protein 5' untranslated region article electroporation epimastigote expression vector gene expression microbial genetics nonhuman priority journal reporter gene Trypanosoma cruzi 5' Untranslated Regions Animals Base Sequence Electroporation Genes, Reporter Genetic Vectors Green Fluorescent Proteins Luciferases Luminescent Proteins Molecular Sequence Data Protozoan Proteins Transfection Trypanosoma cruzi Tubulin Protozoa Trypanosoma Trypanosoma cruzi To improve transfection efficiency in Trypanosoma cruzi, we developed a new electroporation protocol and expression vectors which use luciferase and green and red fluorescent proteins as reporter genes. In transient transfections, the electroporation conditions reported here resulted in luciferase expression 100 times higher than the levels obtained with previously described protocols. To verify whether sequences containing different trans-splicing signals influence reporter gene expression, we compared DNA fragments corresponding to 5′ untranslated plus intergenic (5′ UTR plus Ig) regions from GAPDH, TcP2β, α- and β-tubulin and amastin genes. Vectors containing sequences derived from the first four genes presented similar efficiencies and resulted in luciferase expression in transiently transfected epimastigotes that was up to 10 times higher than that for a control vector. In contrast, the amastin 5′ UTR plus Ig resulted in lower levels of reporter gene expression. We also constructed a vector containing an expression cassette designed to be targeted to the tubulin locus of the parasite. 2004 https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_09320113_v92_n2_p113_DaRocha http://hdl.handle.net/20.500.12110/paper_09320113_v92_n2_p113_DaRocha
institution	Universidad de Buenos Aires
institution_str	I-28
repository_str	R-134
collection	Biblioteca Digital - Facultad de Ciencias Exactas y Naturales (UBA)
topic	alpha tubulin beta tubulin DNA fragment immunoglobulin luciferase red fluorescent protein ribosome protein 5' untranslated region article electroporation epimastigote expression vector gene expression microbial genetics nonhuman priority journal reporter gene Trypanosoma cruzi 5' Untranslated Regions Animals Base Sequence Electroporation Genes, Reporter Genetic Vectors Green Fluorescent Proteins Luciferases Luminescent Proteins Molecular Sequence Data Protozoan Proteins Transfection Trypanosoma cruzi Tubulin Protozoa Trypanosoma Trypanosoma cruzi
spellingShingle	alpha tubulin beta tubulin DNA fragment immunoglobulin luciferase red fluorescent protein ribosome protein 5' untranslated region article electroporation epimastigote expression vector gene expression microbial genetics nonhuman priority journal reporter gene Trypanosoma cruzi 5' Untranslated Regions Animals Base Sequence Electroporation Genes, Reporter Genetic Vectors Green Fluorescent Proteins Luciferases Luminescent Proteins Molecular Sequence Data Protozoan Proteins Transfection Trypanosoma cruzi Tubulin Protozoa Trypanosoma Trypanosoma cruzi Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
topic_facet	alpha tubulin beta tubulin DNA fragment immunoglobulin luciferase red fluorescent protein ribosome protein 5' untranslated region article electroporation epimastigote expression vector gene expression microbial genetics nonhuman priority journal reporter gene Trypanosoma cruzi 5' Untranslated Regions Animals Base Sequence Electroporation Genes, Reporter Genetic Vectors Green Fluorescent Proteins Luciferases Luminescent Proteins Molecular Sequence Data Protozoan Proteins Transfection Trypanosoma cruzi Tubulin Protozoa Trypanosoma Trypanosoma cruzi
description	To improve transfection efficiency in Trypanosoma cruzi, we developed a new electroporation protocol and expression vectors which use luciferase and green and red fluorescent proteins as reporter genes. In transient transfections, the electroporation conditions reported here resulted in luciferase expression 100 times higher than the levels obtained with previously described protocols. To verify whether sequences containing different trans-splicing signals influence reporter gene expression, we compared DNA fragments corresponding to 5′ untranslated plus intergenic (5′ UTR plus Ig) regions from GAPDH, TcP2β, α- and β-tubulin and amastin genes. Vectors containing sequences derived from the first four genes presented similar efficiencies and resulted in luciferase expression in transiently transfected epimastigotes that was up to 10 times higher than that for a control vector. In contrast, the amastin 5′ UTR plus Ig resulted in lower levels of reporter gene expression. We also constructed a vector containing an expression cassette designed to be targeted to the tubulin locus of the parasite.
title	Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
title_short	Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
title_full	Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
title_fullStr	Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
title_full_unstemmed	Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols
title_sort	expression of exogenous genes in trypanosoma cruzi: improving vectors and electroporation protocols
publishDate	2004
url	https://bibliotecadigital.exactas.uba.ar/collection/paper/document/paper_09320113_v92_n2_p113_DaRocha http://hdl.handle.net/20.500.12110/paper_09320113_v92_n2_p113_DaRocha
_version_	1768542512879763456

Expression of exogenous genes in Trypanosoma cruzi: Improving vectors and electroporation protocols

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