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US 5,635,355 AGranted
Methods of obtaining a DNA sequence for integration site independent gene expression
AssigneeMedical Research Council
InventorsFranklin Grosveld, Dimitris Kioussis
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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.
Claims7
A method of obtaining a DNA fragment comprising a dominant activator sequence, comprising
- 1) providing a candidate DNA fragment comprising a DNase I hypersensitive site from a genetic locus containing a structural gene that is expressed in a manner that is specific for a particular mammalian cell type;
- 2) ligating the fragment to an expressible gene to form a construct;
- 3) introducing the construct into the genome of a host cell of said particular mammalian cell type;
- and
- 4) determining if expression of said gene in said particular mammalian cell type is
- (a) 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) independent of the integration site of said construct in said genome,
- wherein such copy number-dependent and integration site-independent gene expression is indicative of the presence of a dominant activator sequence in said candidate DNA fragment.
6 more elements
Dependencies are inferred from the claim text.