Single-copy and amplified CAD genes in Syrian hamster chromosomes localized by a highly sensitive method for in situ hybridization
Syrian hamster cells resistant to N-(phosphonacetyl)-L-aspartate (PALA), a specific inhibitor of the aspartate transcarbamylase activity of the multifunctional protein CAD, overproduce this protein as a result of amplification of the CAD gene. The authors have used a sensitive in situ hybridization technique to localize CAD genes in spreads of metaphase chromosomes from several independent PALA-resistant lines and from wild-type PALA-sensitive cells. The amplified genes were always found within chromosomes, usually in an expanded region of the short arm of chromosome B9. In wild-type cells, the CAD gene was also on the short arm of chromosome B9. In one mutant line, 90 to 100 CAD genes were found within an expanded B9 chromosome and 10 to 15 more were near the distal end of one arm of several different chromosomes. Another line contained most of the genes in a telomeric chromosome or large chromosome fragment. The amplified genes were in chromosomal regions that were stained in a banded pattern by trypsin-Giemsa. A few double minute chromosomes were observed in a very small fraction of the total spreads examined. The in situ hybridizations were performed in the presence of 10% dextran sulfate 500, which increases the signal by as much as 100-fold. Using recombinant DNA plasmids nick-translated with -/sup 125/I)dCTP to high specific radioactivity, 10 CAD genes in a single chromosomal region were revealed after 1 week of autoradiographic exposure, and the position of the unique gene could be seen after 1 month.
- Research Organization:
- Tumor Virology Lab., The Salk Institute, La Jolla, CA 92037
- OSTI ID:
- 5652951
- Journal Information:
- Mol. Cell. Biol.; (United States), Vol. 2:3
- Country of Publication:
- United States
- Language:
- English
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Related Subjects
TRANSFORMATIONS
GENE AMPLIFICATION
GENETIC MAPPING
TRANSPOSONS
ASPARTIC ACID
AUTORADIOGRAPHY
CELL CULTURES
CHROMOSOMAL ABERRATIONS
CHROMOSOMES
CLONE CELLS
ENZYME INHIBITORS
GENES
HAMSTERS
HYBRIDIZATION
IODINE 125
MUTANTS
PLASMIDS
RECOMBINANT DNA
SENSITIVITY
TECHNOLOGY ASSESSMENT
AMINO ACIDS
ANIMALS
BETA DECAY RADIOISOTOPES
CARBOXYLIC ACIDS
CELL CONSTITUENTS
DAYS LIVING RADIOISOTOPES
DNA
ELECTRON CAPTURE RADIOISOTOPES
INTERMEDIATE MASS NUCLEI
IODINE ISOTOPES
ISOTOPES
MAMMALS
MAPPING
MUTATIONS
NUCLEI
NUCLEIC ACIDS
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
RADIOISOTOPES
RODENTS
VERTEBRATES
550401* - Genetics- Tracer Techniques
550201 - Biochemistry- Tracer Techniques