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US 5,849,997 AGranted
Vector for integration site independent gene expression in mammalian host cells
AssigneeMedical Research Council
InventorsFranklin Grosveld, Dimitris Kioussis
Life of this patent
Estimating the term…
Abstract
Vectors for the integration of a gene into the genetic material of a mammalian host cell such that the gene may be expressed by the host cell comprise a promoter and the gene and include a dominant activator sequence capable of eliciting host cell-type restricted, integration site independent, copy number dependent expression of the gene.
Claims22
A non-human transgenic mammal comprising, integrated into its genome, a recombinant DNA comprising:
- (i) a dominant acitivator sequence that is specific for a particular mammal cell-type,
- (ii) a promoter that is functional in said particular mammal cell-type, and
- (iii) a structural gene operatively linked to said promoter, wherein the region in said DNA consisting of (i), (ii) and (iii) and DNA therebetween has a nuclrotide sequence different from that of a naturally occurring DNA sequence, and said dominant activator sequence being characterized in that:
- (I) it is associated in naturally occurring DNA with a naturally occurring gene that is expressed in a tissue-specific manner;
- and
- (II) it is locatable in naturally occurring DNA by association with a DNase I super hypersensitive site;
- wherein said recombinant DNA is integrated into a genomic locus that is different from the genomic locus of the dominant activator sequence in naturally occurring DNA and wherein said dominant activator sequence stimulated expression of said structural gene when said DNA molecule is integrated into a genome of a host cell of said cell-type, such that said expression:
- (a) is dependent on the number of copies of said gene that are integrated into said genome in that said expression increases as said number of copies of said gene increases;
- (b) is independent of the integration site of said DNA molecule in said genome;
- and
- (c) results in production in said host cell of said cell type of a detectable amount of product encoded by said structural gene.
8 more elements
A non-human transgenic mammal of a first species comprising, integrated into its genome, a recombinant DNA comprising:
- (i) a dominant activator sequence from a second species of mammals that is specific for a particular mammal cell-type;
- (ii) a promoter that is functional in said particular mammal cell-type, and
- (iii) a structural gene operatively linked to said promoter, wherein the region in said DNA consisting of (i), (ii) and DNA therebetween has a nucleotide sequence different from that of a naturally occurring DNA sequence, and said dominant activator sequence being characterized in that:
- (I) it is associated in naturally occurring DNA with a naturally occurring gene that is expressed in a tissue-specific manner;
- and
- (II) it is locatable in naturally occurring DNA by association with a DNase I super hypersensitive site;
- wherein said dominant activator sequence stimulated expression of said structural gene when said DNA molecule is integrated into a genome, of a host cell of said cell-type, such that said expression:
- (a) is dependent on the number of copies of said gene that are integrated into said genome in that said expression increases as said number of copies of said gene increases;
- (b) is independent of the integration site of said DNA molecule in said genome;
- and
- (c) results in production in said host cell of said cell type of a detectable amount of product encoded by said structural gene.
8 more elements
A method of obtaining production of a protein encoded by a structural gene in a particular cell-type of a transgenic animal comprising
- A) providing a transgenic animal containing a recombinant DNA comprising:
- (i) a dominant activator sequence that is specific for a particular cell-type,
- (ii) a promoter that is functional in said particular cell-type, and
- (iii) said structural gene operatively linked to said promoter, wherein the region in said DNA consisting of (i), (ii) and (iii) and DNA therebetween has a nucleotide sequence different from that of naturally occurring DNA sequence, and said dominant activator sequence being characterized in that,
- (I) it is associated in naturally occurring DNA with a naturally occurring gene that is expressed in a tissue-specific manner, and
- (II) it is locatable in naturally occurring DNA by association with a DNase I super hypersensitive site;
- wherein said dominant activator sequence is characterized in that it stimulates expression of said structural gene where said DNA molecule is integrated into a genome of a host cell of said cell-type, such that said expression:
- (a) is dependent on the number of copies of said gene that are integrated into said genome in that said expression increases as said number of copies of said gene increases;
- and
- (b) is independent of the integration site of said DNA molecule in said genome;
- and
- B) permitting expression of said structural gene to produce said protein in said cell-type.
9 more elements
A method of obtaining a transgenic animal containing, integrated into its genome, a structural gene that is expressed in a particular cell-type of said animal the method comprising introducing into the genome of a animal a recombinant DNA comprising
- (i) a dominant activator sequence that is specific for a particular animal cell-type,
- (ii) a promoter that is functional in said particular animal cell-type, and
- (iii) a structural gene operatively linked to said promoter, wherein the region in said DNA consisting of (i), (ii) and (iii) and DNA therebetween has a nucleotide sequence different from that of naturally occurring DNA, and said dominant activator sequence being characterized in that in naturally occurring DNA:
- (I) it is associated with a naturally occurring gene that is expressed in a tissue-specific manner;
- and
- (II) it is locatable in naturally occurring DNA by association with a DNase I super hypersensitive site;
- wherein said dominant activator sequence is characterized in that it stimulates expression of said structural gene when said DNA molecule is integrated into a genome of a host cell of said cell-type, such that said expression:
- (a) is dependent on the number of copies of said gene that are integrated into said genome in that said expression increases as said number of copies of said gene increases;
- (b) is independent of the integration site of said DNA molecule in said genome;
- and
- (c) results in production in said host cell type of a detectable amount of product encoded by said structural gene.
8 more elements
Dependencies are inferred from the claim text.