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Doctoral Thesis
DOI
https://doi.org/10.11606/T.87.2013.tde-16052014-092549
Document
Author
Full name
Lucas Martins Chaible
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2013
Supervisor
Committee
Dagli, Maria Lucia Zaidan (President)
Belizario, Jose Ernesto
Chiaratti, Marcos Roberto
Meirelles, Flavio Vieira
Strauss, Bryan Eric
Title in Portuguese
Criação e caracterização de um modelo transgênico inédito de camundongos com expressão condicional do gene da conexina 43.
Keywords in Portuguese
Camundongos
Conexinas
Expressão gênica
Genótipo
Transgenes
Abstract in Portuguese
As conexinas (Cx) compõem as junções comunicantes do tipo gap, entre elas a Cx43 é a mais prevalente. A diminuição de sua expressão está relacionada com diversas alterações fisiológicas. Sua importância in vivo foi relatada em camundongos com deleção de um dos alelos de Cx43 (Cx43+/-), pois os animais Cx43-/- não são viáveis. Devido esta inviabilidade técnica, os estudos com essa proteína foram realizados com animais Cx43+/-. Assim, este trabalho propõe a criação de um modelo transgênico para o estudo da Cx43. Para isso, o gene Cx43 foi reintroduzido ao genoma murino, porém regulado pelo sistema induzido por doxiciclina TetOn. Vetores de expressão gênica foram construídos e validados quanto sua funcionalidade in vitro e posterior transferencia para zigotos murinos por meio de microinjeção pronuclear. Obtivemos sucesso na construção do sistema de expressão. A funcionalidade dos vetores foi confirmada in vitro utilizando células HeLa e E10. Nos experimentos in vivo, apesar das taxas adequadas de nascimento nenhum deles apresentou genótipo positivo para o transgene.
Title in English
Establishment and characterization of a novel transgenic mouse model with conditional expression of connexin 43 gene.
Keywords in English
Connexins
Gene expression
Genotype
Mice
Transgenes
Abstract in English
The connexins (Cx) comprise gap junctions type, and the Cx43 is the most prevalent. The decrease of its expression is related to various physiological changes. Its importance in vivo has been reported in mice with deletion of one allele of Cx43 (Cx43+/-), because the animals Cx43-/- are not viable. So all studies with this protein were performed with animals Cx43+/-. Thus, this paper proposes the creation of new a transgenic model for the study of Cx43 protein. For this, the Cx43 gene was reintegrated to the murine genome, but drived by doxycycline Teton inducible system. Gene expression vectors were constructed and validated in vitro and subsequent transfer to mouse zygotes by pronuclear microinjection. We got success in vector construction and functionality. The functionality of the vectors was confirmed in vitro using HeLa cells and E10. In experiments in vivo, despite adequate rates of birth, no one animal showed positive genotype for the transgene.
 
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Publishing Date
2014-05-21
 
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