DNA sequence which acts as a chromatin insulator element to protect expressed genes from cis-acting regulatory sequences in mammalian cells

An isolated DNA construct for insulating the differential expression of two genes, comprising:

• (a) one or more isolated eukaryotic insulator molecules according to any of 1, 2, or 3;
• (b) a first expressible gene;
• (c) a second expressible gene;
• (d) a promoter that mediates expression of said first gene operably linked to said first gene;
• (e) a promoter that mediates expression of said second gene operably linked to said second gene;
• (f) an enhancer operably linked to the second gene such that said enhancer enhances expression of said second gene relative to the expression of said second gene lacking said operably linked enhancer;
• wherein said one or more of the insulator molecules is positioned in the construct 5' of the promoter operably linked to the first gene;
• wherein the enhancer is positioned 5' of the promoter operably linked to the second gene which is positioned in opposite transcriptional orientation to said first gene;
• and further wherein one or more of the insulators is positioned 3' of the first and second genes.

An isolated DNA construct for incorporation into a host cells and for insulation of the expression of a gene therein, comprising:

• (a) DNA comprising a transcription unit comprising an heterologous gene, a promoter to drive transcription of the gene, and an enhancer element;
• and
• (b) one or more insulator elements having insulator activity and operative in eukaryotic cells, wherein said isolated insulator DNA element consists of a eukaryotic 5' constitutive DNase I-hypersensitive site from the 5' region of the chicken .beta.-globin gene locus and is isolated from a 1.2 kilobase SacI-SspI DNA fragment and minimally comprises the DNA of Seq. ID. No.1, and further wherein said insulator element is positioned in sufficient proximity to the transcription unit to insulate the transcription and expression of the gene from cis-acting DNA regulatory sequences in chromatin into which the gene has integrated;
• said insulator element being outside of the DNA according to (a) above.

An isolated DNA construct for incorporation into a host cells and for insulation of the expression of a gene therein, comprising:

• (a) DNA comprising a transcription unit comprising an heterologous gene and a promoter to drive transcription of the gene;
• and
• (b) one or more insulator elements having insulator activity and operative in eukaryotic cells, wherein said isolated insulator DNA element consists of a eukaryotic 5' constitutive DNase I-hypersensitive site from the 5' region of the chicken .beta.-globin gene locus and is isolated from a 1.2 kilobase SacI-SspI DNA fragment and minimally comprises the DNA of Seq. ID. No.1, and further wherein said insulator element is positioned in sufficient proximity to the transcription unit to insulate the transcription and expression of the gene from cis-acting DNA regulatory sequences in chromatin into which the gene has integrated;
• said insulator element being outside of the DNA according to (a) above.