Candida utilis transformation system

C - Chemistry – Metallurgy – 12 – N

Patent

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Details

C12N 15/81 (2006.01) C12N 9/24 (2006.01) C12N 9/26 (2006.01) C12N 9/88 (2006.01)

Patent

CA 2268004

The present invention discloses a transformation system useful to express heterologous proteins in the Candida utilis yeast, based on obtaining auxotrophic mutants of said species as well as the isolation of different genes, from a genomic library, which complement said auxotrophies. The transformation system uses as hosts new auxotrophic mutants obtained from the yeast NRRL Y-1084 of Candida utilis which are defective mainly in the biosynthetic ways of uracyl and histidine, which are transformed with plasmids containing as selection markers the genes URA3 and HIS3 of Candida utilis. Another aspect of the invention is the isolation of the gene coding for the enzyme sucrose invertase or .beta.-fructofuranosidase of Candida utilis, as well as the identification of sequences for promoting, secretion signalling and termination of said gene INV1. These sequences are useful to obtain the expression of heterologous proteins in said yeast.

L'invention concerne un système de transformation utile pour exprimer des protéines hétérologues dans la levure Candida utilis, ce système de transformation se fondant sur l'obtention de mutants auxotrophiques de cette espèce ainsi que sur l'isolement de différents gènes, à partir d'une bibliothèque génomique, qui complètent lesdites auxotrophies. Le système de transformation consiste à utiliser en tant que hôtes de nouveaux mutants auxotrophiques obtenus à partir de la levure NRRL Y-1084 de Candida utilis, lesquels sont déficients essentiellement dans les voies biosynthétiques de uracyl et histidine, lesquels sont transformés avec des plasmides qui contiennent en tant que marqueurs de sélection les gènes $(URA3) et HIS3 de $(Candida utilis). Un autre aspect de l'invention concerne l'isolement du gène codant pour l'enzyme sucrose invertase ou .beta.-fructofuranosidase de Candida utilis, ainsi que l'identification de séquences de promotion, de signalisation de sécrétion et de terminaison dudit gène INV1. Ces séquences peuvent être utilisées de manière avantageuse pour effectuer l'expression de protéines hétérologues dans cette levure.

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