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1. WO2001062944 - INTERGENIC AND INTRAGENIC INTEGRATION SITES FOR FOREIGN GENE EXPRESSION IN RECOMBINANT S. GORDONII STRAINS

Publication Number WO/2001/062944
Publication Date 30.08.2001
International Application No. PCT/US2001/005493
International Filing Date 22.02.2001
Chapter 2 Demand Filed 21.09.2001
IPC
C12N 15/74 2006.1
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
74Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
CPC
C12N 15/74
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
74Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
C12N 15/746
CCHEMISTRY; METALLURGY
12BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
NMICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
15Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
09Recombinant DNA-technology
63Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
74Vectors or expression systems specially adapted for prokaryotic hosts other than E. coli, e.g. Lactobacillus, Micromonospora
746for lactic acid bacteria (Streptococcus; Lactococcus; Lactobacillus; Pediococcus; Enterococcus; Leuconostoc; Propionibacterium; Bifidobacterium; Sporolactobacillus)
Applicants
  • SIGA PHARMACEUTICALS, INC. [US]/[US] (AllExceptUS)
  • FRANKE, Christine, A. [US]/[US] (UsOnly)
  • BOLKEN, Tove, C. [US]/[US] (UsOnly)
  • JONES, Kevin, F. [US]/[US] (UsOnly)
  • HRUBY, Dennis, E. [US]/[US] (UsOnly)
Inventors
  • FRANKE, Christine, A.
  • BOLKEN, Tove, C.
  • JONES, Kevin, F.
  • HRUBY, Dennis, E.
Agents
  • REA, Teresa, Stanek
Priority Data
60/184,64524.02.2000US
Publication Language English (en)
Filing Language English (EN)
Designated States
Title
(EN) INTERGENIC AND INTRAGENIC INTEGRATION SITES FOR FOREIGN GENE EXPRESSION IN RECOMBINANT S. GORDONII STRAINS
(FR) NOUVEAUX SITES D'INTEGRATION INTERGENIQUE ET INTRAGENIQUE POUR L'EXPRESSION DE GENES ETRANGERS DANS DES SOUCHES S. GORDONII RECOMBINANTES
Abstract
(EN) The present invention provides two new chromosomal integration sites for expression of foreign genes have been developed in Streptococcus gordonii (S. gordonii). One integration site is intergenic between orfA and orfB in an operon of unknown function. The other site is intragenic within the lacG gene, which encodes phospho- $g(b)-galactosidase, and is part of the lactose (lac) operon. Them6 gene from Streptococcus pyogenes was integrated in a stable configuration into the chromosome of S. gordonii at each of these integration sites, and in both cases the recombinant bacteria expressed the M6 protein on their surface. Furthermore, expression from the lacG site within the lactose operon was shown to be regulated by growth on lactose. Identification of these new chromosomal insertion sites provides the ability to express multiple foreign genes from the same recombinant and the potential for modulating expression in vitro or in vivo by the use of a biosynthetic metabolite.
(FR) La présente invention concerne deux nouveaux sites d'intégration de chromosomes destinés à l'expression de gènes étranges; ils ont été développées dans Streptococcus gordonii (S. gordonii). Un site d'intégration est intergénique entre orfA et orfB, dans un opéron à fonction inconnue. L'autre site est intragénique et se situe à l'intérieur du gène lacG qui code phospho- $g(b)-galactosidase; il fait partie de l'opéron lactose (lac). Le gène emm6 provenant de Streptococcus pyogenes a été intégré dans une configuration stable dans le chromosome de S. gordonii dans chacun de ces sites d'intégration; dans les deux cas, les bactéries recombinantes exprimaient la protéine M6 à leur surface. En outre, on a montré que l'expression partant du site lacG à l'intérieur de l'opéron de lactose a été régulée par la croissance sur le lactose. L'identification de ces nouveaux sites d'insertion de chromosomes permet d'exprimer des gènes étrangers multiples à partir de la même recombinante et offrent une possibilité de moduler l'expression in vitro ou in vivo par l'utilisation d'un métabolite biosynthétique.
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