Abstract
We have generated mice having a single copy of the human haptoglobin gene (Hp2), driven by its natural promoter, and a neomycin resistance gene (Neo), driven by a herpes simplex thymidine kinase promoter with polyoma enhancers, inserted into two defined chromosomal locations, the Hprt locus on the X-chromosome and the apolipoprotein (apo) AI-CIII gene cluster on chromosome 9. The haptoglobin promoter is highly specialized in its tissue of action; the viral promoter has few restrictions. The apoAI-CIII gene is naturally active in only two tissues, whereas the Hprt gene region is ubiquitously active. Expression of both transgenes at substantial levels was achieved only (a) when the transgenes were inserted into the genome close to a known tissue-specific enhancer/locus control region in the apoAI-CIII gene cluster, and (b) when known conditions for function of their promoters were met. The specificities of the two chromosomal regions and of the two promoters are preserved, but their interactions are not specific. We conclude that transgenes are affected by locus-related enhancers in the same manner as nearby endogenous genes. Our experiments reinforce the usefulness of using gene targeting to direct single-copy transgenes to appropriate chromosomal locations.
Highlights
Transgenic mouse technology has been widely and effectively used for studying the physiological consequences of expressing exogenous gene products or analyzing promoter function and protein structure-function relationships in vivo [1]
Our results show that the chromosomal location of a transgene markedly and with a considerable degree of predictability affects the levels of its expression
neomycin resistance gene (Neo) gene in the apoAI-CIII region of the Apo-Hp2 mice takes on the same strong expression pattern as the apolipoprotein locus, which is highly expressed in the liver and intestine
Summary
The Neo-hHp2 Transgene Cassette—The entire human Hp2 gene [10] is contained within a 9-kb fragment extending from an XbaI site at approximately 1.1 kb 5Ј to the starting codon to an ApaI site 1.6 kb 3Ј to the stop codon. A 9-kb BamHI fragment of mouse genomic DNA containing the Apoa-1 gene and a 3Ј part of the Apoc-3 gene [7] was used to make the targeting construct designed to insert the Neo-hHp2 cassette into the apoAI-CIII region. The human haptoglobin  probe was a 650-bp BamHI-HindIII fragment corresponding to exon 7 of the Hp2 gene. The resulting radioactive RNA was hybridized to immobilized DNA probes for apoC-III, apoA-I, Neo, and Gapdh described above
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