C - Chemistry – Metallurgy – 12 – N
Patent
C - Chemistry, Metallurgy
12
N
C12N 1/21 (2006.01) A61K 39/112 (2006.01) C12N 15/11 (2006.01)
Patent
CA 2706204
- 52 - Method for producing multiple modifications in the chromosome of Gram- negative bacteria and Salmonella strains which are deficient in c-di-GMP synthesis obtained by said method. The method can be used to make multiple modifications in the genome of Gram-negative bacteria, simply and efficiently, using the plasmids of the invention, which comprise a marker gene under the control of a constitutive promoter, a replication origin specific for Gram- negative bacteria, a gene which encodes a heat-sensitive protein essential for initiating replication of the plasmid, making the replication origin heat-sensitive, and a counter-selection gene. The invention also relates to mutant Salmonella enterica strains obtained using the method of the invention in which some of the twelve genes which encode proteins with GGDEF domain have been selected*, and to the use thereof as expression vectors, immunotherapeutic agents and in metabolic studies
Procedimiento que permite producir modificaciones múltiples en el cromosoma de bacterias gram negativas y cepas de Salmonella deficientes en síntesis de c-di-GMP obtenidas por el mismo. El procedimiento permite realizar múltiples modificaciones en el genoma de bacterias Gram negativas de forma sencilla y eficaz, utilizando los plásmidos de la invención, que contienen un gen indicador bajo el control de un promotor constitutivo, un origen de replicación específico para bacterias Gram negativas, un gen que codifica una proteína termosensible esencial para la iniciación de la replicación del plásmido, convirtiendo el origen de replicación en termosensible, y un gen de contra-selección. La invención se refiere también a cepas mutantes de Salmonella enterica obtenidas mediante el procedimiento de la invención en las que parte o los doce genes que codifican proteínas con dominio GGDEF han sido delecionadosy al uso de las mismas como vectores de expresión, agentes inmunoterapéuticos y en estudios metabólicos.
Garcia Begona
Lasa Inigo
Latasa Cristina
Solano Cristina
Toledo-Arana Alejandro
Bereskin & Parr Llp/s.e.n.c.r.l.,s.r.l.
Consejo Superior de Investigaciones Cientificas
Universidad Publica de Navarra
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