Method for producing a marker-free mutated target organism and plasmid vectors suitable for the same
OwnerMarkus Pompejus, Corinna Klopprogge +2inventors
InventorsMarkus Pompejus, Corinna Klopprogge, Oskar Zelder, Wolfgang Liebl
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Abstract
The invention relates to a plasmid vector which does not replicate in a target organism, comprising the following components: a) an origin of replication for a host organism which is different from the target organism, b) at least one genetic marker, c) where appropriate, a sequence section which makes possible the transfer of DNA via conjugation (mob sequence), d) a sequence section which is homologous to sequences of the target organism and makes possible homologous recombination in the target organism, e) a gene for a galactokinase under the control of a promotor.
Claims15
A plasmid vector which does not replicate in a target organism, comprising the following components:
- a) an origin of replication for a host organism which is different from the target organism,
- b) at least one genetic marker,
- c) where appropriate, a sequence section which makes possible the transfer of DNA via conjugation (mob sequence),
- d) a sequence section which is homologous to sequences of the target organism and makes possible homologous recombination in the target organism,
- e) a gene for a galactokinase under the control of a promotor.
A method for preparing a marker-free mutated target organism, comprising the following steps:
- a) transferring a plasmid vector as claimed in any of claims 1 to 10 into a target organism,
- b) selecting clones of said target organism, which contain at least one genetic marker introduced by said plasmid vector,
- c) selecting the clones of said target organism, obtained in step b), for the presence of galactose sensitivity by culturing in a galactose-containing medium.
The use of a galactokinase gene as conditionally negatively dominant marker gene.
Dependencies are inferred from the claim text.