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. 1990 Dec;10(12):6619–6623. doi: 10.1128/mcb.10.12.6619

Functional expression of the genomic DNA sequences encoding mouse Na,K-ATPase alpha 1 gene by cotransfection of overlapping genomic DNA segments.

S Y Tam 1, E N Geissler 1, S L Graw 1, D E Housman 1
PMCID: PMC362939  PMID: 2174112

Abstract

The entire 33-kb coding region of the mouse Na,K-ATPase alpha 1 subunit gene was cloned in two overlapping cosmids which contain inserts of 40 kb. To assess the functional expression of the mouse alpha 1 gene, the two cosmids were cotransfected into ouabain-sensitive CV-1 monkey cells yielding an average of 64 resistant colonies per 10(6) cells per microgram of DNA. Analysis of the DNA transferred to the ouabain-resistant transformants by the two cosmids suggests that the generation of a functional gene can occur by homologous recombination between the two introduced segments, as demonstrated by generation of a novel diagnostic restriction fragment. The ability to reconstruct the intact mouse alpha 1 gene in a heterologous host cell and to monitor its functional expression with a selection protocol permits direct identification and isolation of regulatory sequences for the gene.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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