Titre :
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Improvement of a non-viral gene transfer strategy : Obtaining of hyperactive mutants of Mos1 Mariner transposase (abstract : congrès international de Myologie, 2005)
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contenu dans :
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Auteurs :
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Congrès international de myologie 2005 (International Congress of Myology 2005; 9-13 mai 2005; Nantes, France) ;
Germon S ;
Augé-Gouillou C ;
Bonnin-Rouleux F ;
Brillet B ;
Bigot Y
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Type de document :
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Article
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Année de publication :
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2005
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Pages :
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p. 277
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Langues:
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Anglais
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Mots-clés :
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ARN messager
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colloque
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encapsulation
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mutation génétique
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plasmide
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thérapie génique
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vecteur non viral
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Résumé :
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Communication n° 154 Improvement of a non-viral gene transfer strategy: Obtaining of hyperactive mutants of Mos1 Mariner transposase. S Germon, C Augé-Gouillou, F Bonnin-Rouleux, B Brillet and Y Bigot. Laboratoire d'Etude des Parasites Génétiques. Faculté des Sciences et Techniques. Avenue Monge 37200 Tours - France. Introduction : We are developing a synthetic gene transfer strategy based on the use of Mos-1 Mariner transposon. Such element consists in a unique open reading frame - encoding for a transposase - flanked by two inverted terminal sequences (ITR). The transposase is necessary and sufficient for the transposition by a "cut and paste" mechanism. For gene transfer, the gene of interest could be placed between the two ITR and the transposase supplied in trans via either an expression plasmid or mRNA. Synthetic gene transfer strategies offer a safe process but present a low efficiency mainly due to a decreased nuclear import. The use of large transgene quantity could lead to a higher rate of non-homologous recombination and transposase-dependant recombination. Objectives : In order to improve the efficiency of this strategy, we are investigating several options and particularly the obtaining of more efficient Mariner transposases. Methods : Some specific residues (10) were mutated by site-directed mutagenesis. Transposition efficiency of these mutants was evaluated with a transposition test in bacteria (antibiotic resistance acquired during the transposition event) and the most promising mutations were combined to study the synergy or the antagonism of the mutations. Results : Single mutations lead to an increased transposition activity (10 to 17-fold) and some multiple mutations allow us to obtain a transposase up to 60-fold more active. Conclusions : We were able to obtain some hyperactive transposase mutants in bacteria and we are now evaluating their efficiency in mammalian cells.
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