A system that selects for the gene directing synthesis of the enzyme adenine phosphoribosyltransferase (APRT) uses the antibiotic alanosine to prevent endogenous synthesis of adenylic acid. With the aid of this system, a new series of human-mouse hybrids has been prepared between wild type human diploid fibroblasts and an enzyme-deficient mouse line. Survival of the hybrids depended upon the presence of the APRT, which was shown to have the isoelectric pH characteristic of the human enzyme and not that of the mouse. Reduced hybrids containing the enzyme lacked all human biarmed chromosomes, so that unless a rearrangement had occurred, the aprt gene must be located on an acrocentric chromosome. The hybrid cells became APRT-with a frequency of 2 X 10-3, probably by loss of the human aprt chromosome. The APRT-progeny could be obtained selectively by growth in medium containing fluoroadenine.Human-mouse hybrid cells eliminate human chromosomes on serial culture (1), but in a suitably constituted hybrid (2) a chemical selective agent may make a human gene necessary for viability. The hybrid cells need therefore retain only a single human chromosome (3) or portion thereof (4) that bears the essential gene. Demonstration of this for the human thyridine kinase gene (3, 4) permitted an assignment of this gene to an E group chromosome (no. 17 or 18).In order to generalize such a method it is desirable to introduce an enzymatic deficiency into each of a number of mouse lines, make hybrids with human diploid cells, and reduce the complements of human chromosomes to single chromosomes (5). We now describe a selective system for the enzyme adenine phosphoribosyl transferase (APRT) and the production of reduced mouse-human hybrids containing the human enzyme.
MATERIALS AND METHODSThe standard growth medium was that of Dulbecco and Vogt, equilibrated with 10% CO2-90% air.
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