Mutation at the adenine phosphoribosyl transferase locus occurs at a higher than expected frequency in both Chinese hamster ovary cells and mouse L-cells. As a result of 2D gel analysis of mutant proteins from both heterozygotes and homozygotes a model is presented hypothesizing a two step mechanism for the production of mutants in eukaryotic cells, one step of which involves gene inactivation. The mechanism of gene inactivation will be studied using a series of DNA probes constructed from a CH-APRT gene pBR322 clone. Wild type APRT DNA will be cloned into bacteriophage M13, and this phage used as a template for the isolation of mutant DNA. Using the M13 dideoxy-system we shall sequence mutant and wild-type DNA. This will be accompanied by S1-nuclease digestion of hybrid DNA to locate alterations in DNA sequences. The E. coli apt gene has been cloned, and will be used to transform APRT- strains of mammalian cells. These can then be used to study transformation and in vitro mutagenesis. HPLC analysis of peptides from human and rodent APRT will continue.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK025498-07
Application #
3227434
Study Section
Mammalian Genetics Study Section (MGN)
Project Start
1980-01-01
Project End
1987-12-31
Budget Start
1986-01-01
Budget End
1986-12-31
Support Year
7
Fiscal Year
1986
Total Cost
Indirect Cost
Name
Indiana University Bloomington
Department
Type
Schools of Arts and Sciences
DUNS #
006046700
City
Bloomington
State
IN
Country
United States
Zip Code
47402