Single-copy and amplified CAD genes in Syrian hamster chromosomes localized by a highly sensitive method for in situ hybridization

Journal Article · Mon Mar 01 04:00:00 EST 1982 · Mol. Cell. Biol.; (United States)

DOI:https://doi.org/10.1128/MCB.2.3.308· OSTI ID:5652951

Wahl, G M; Vitto, L; Padgett, R A; Stark, G R

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), Journal Name: Mol. Cell. Biol.; (United States) Vol. 2:3; ISSN MCEBD
• Country of Publication: United States
• Language: English

Related Subjects

550201 -- Biochemistry-- Tracer Techniques
550401* -- Genetics-- Tracer Techniques
59 BASIC BIOLOGICAL SCIENCES
AMINO ACIDS
ANIMALS
ASPARTIC ACID
AUTORADIOGRAPHY
BETA DECAY RADIOISOTOPES
CARBOXYLIC ACIDS
CELL CONSTITUENTS
CELL CULTURES
CHROMOSOMAL ABERRATIONS
CHROMOSOMES
CLONE CELLS
DAYS LIVING RADIOISOTOPES
DNA
ELECTRON CAPTURE RADIOISOTOPES
ENZYME INHIBITORS
GENE AMPLIFICATION
GENES
GENETIC MAPPING
HAMSTERS
HYBRIDIZATION
INTERMEDIATE MASS NUCLEI
IODINE 125
IODINE ISOTOPES
ISOTOPES
MAMMALS
MAPPING
MUTANTS
MUTATIONS
NUCLEI
NUCLEIC ACIDS
ODD-EVEN NUCLEI
ORGANIC ACIDS
ORGANIC COMPOUNDS
PLASMIDS
RADIOISOTOPES
RECOMBINANT DNA
RODENTS
SENSITIVITY
TECHNOLOGY ASSESSMENT
TRANSFORMATIONS
TRANSPOSONS
VERTEBRATES