Efficient procedure for transferring specific human genes into Chinese hamster cell mutants: interspecific transfer of the human genes encoding leucyl- and asparaginyl-tRNA synthetases

Journal Article · Sun May 01 04:00:00 EDT 1983 · Mol. Cell. Biol.; (United States)

DOI:https://doi.org/10.1128/MCB.3.5.892· OSTI ID:6748647

Cirullo, R E; Dana, S; Wasmuth, J J

A simple and efficient procedure for transferring specific human genes into mutant Chinese hamster ovary cell recipients has been developed that does not rely on using calcium phosphate-precipitated high-molecular-weight DNA. Interspecific cell hybrids between human leukocytes and temperature-sensitive Chinese hamster cell mutants with either a thermolabile leucyl-tRNA synthetase or a thermolabile asparaginyl-tRNA synthetase were used as the starting material in these experiments. These hybrids contain only one or a few human chromosomes and require expression of the appropriate human aminoacyl-tRNA synthetase gene to grow at 39 degrees C. Hybrids were exposed to very high doses of gamma-irradiation to extensively fragment the chromosomes and re-fused immediately to the original temperature-sensitive Chinese hamster mutant, and secondary hybrids were isolated at 39 degrees C. Secondary hybrids, which had retained small fragments of the human genome containing the selected gene, were subjected to another round of irradiation, refusion, and selection at 39 degrees C to reduce the amount of human DNA even further. Using this procedure, Chinese hamster cell lines have been constructed that express the human genes encoding either asparaginyl- or leucyl-tRNA synthetase, yet less than 0.1% of their DNA is derived from the human genome, as quantitated by a sensitive dot-blot nucleic acid hybridization procedure.

• Research Organization: Univ. of California, Irvine
• OSTI ID: 6748647
• Journal Information: Mol. Cell. Biol.; (United States), Journal Name: Mol. Cell. Biol.; (United States) Vol. 3:5; ISSN MCEBD
• Country of Publication: United States
• Language: English

Related Subjects

550400 -- Genetics
560121* -- Radiation Effects on Cells-- External Source-- (-1987)
59 BASIC BIOLOGICAL SCIENCES
63 RADIATION, THERMAL, AND OTHER ENVIRON. POLLUTANT EFFECTS ON LIVING ORGS. AND BIOL. MAT.
ANIMAL CELLS
ANIMALS
BIOLOGICAL EFFECTS
BIOLOGICAL MATERIALS
BIOLOGICAL RADIATION EFFECTS
BLOOD
BLOOD CELLS
BODY FLUIDS
CHROMOSOMES
CLONING
CONNECTIVE TISSUE CELLS
DNA-CLONING
ELECTROMAGNETIC RADIATION
ENZYMES
FIBROBLASTS
GAMMA RADIATION
GENE RECOMBINATION
GENETIC EFFECTS
GENETIC RADIATION EFFECTS
HAMSTERS
HYBRIDIZATION
IONIZING RADIATIONS
LEUKOCYTES
LIGASES
MAMMALS
MAN
MATERIALS
MUTANTS
NUCLEIC ACIDS
ORGANIC COMPOUNDS
PRIMATES
RADIATION EFFECTS
RADIATIONS
RNA
RODENTS
SOMATIC CELLS
TRANSFER RNA
VERTEBRATES