Effects of flanking base sequences on 5-bromodeoxyuridine mutagenesis in mammalian cells
| Autor: | Mark T. Kresnak, Richard L. Davidson |
|---|---|
| Rok vydání: | 1991 |
| Předmět: |
Alkylating Agents
Hypoxanthine Phosphoribosyltransferase Guanine Mutant Molecular Sequence Data Mutagenesis (molecular biology technique) Biology DNA sequencing Cell Line chemistry.chemical_compound Mice Shuttle vector Genetics Animals Gene Base Composition Base Sequence Nucleic acid sequence Methane sulfonate Cell Biology General Medicine Molecular biology chemistry Biochemistry Bromodeoxyuridine Gene Expression Regulation Mutagenesis Ethyl Methanesulfonate Mutagens |
| Zdroj: | Somatic cell and molecular genetics. 17(4) |
| ISSN: | 0740-7750 |
| Popis: | The molecular mechanisms of incorporation-dependent, 5-bromodeoxyuridine (BrdU)-induced mutagenesis were analyzed in murine A9 cells that possess a single copy of the Escherichia coli gpt gene integrated into the chromosomal DNA as part of a shuttle vector. Four independently derived GPT- mutants with single base changes within the integrated gpt gene were utilized in BrdU-induced reversion analyses to test the relative mutability of guanine residues in four different settings: the 5' and 3' guanine residues of a GG doublet, the 3' guanine residue of a GGGG quartet, and the middle guanine residue of a GGG triplet. Two of the mutant lines possessed GG doublet sequences in which a GC----AT transition at either guanine residue of the doublet leads to restoration of GPT enzyme activity without restoring wild-type DNA sequence. Both lines were shown to be effectively reverted by BrdU incorporation-dependent mutagenesis, and sequencing of the gpt genes from numerous independently derived revertants of both lines demonstrated that greater than 90% of the revertants arose due to GC----AT transitions at the 3' guanine residue of the doublet. BrdU-induced reversion of two additional GPT- mutant lines demonstrated that the 3' guanine residue of a GGGG quartet is efficiently mutated, while the middle guanine residue of a GGG triplet sequence is at least 10-fold less mutable by BrdU incorporation-dependent mutagenesis than the 3' guanine residue of a GG doublet or GGGG quartet. All four mutant lines tested were equally revertible by treatment with the alkylating agent ethyl methane sulfonate. The results from this study define a sequence-specific mechanism for BrdU-induced, incorporation-dependent mutagenesis and demonstrate the use of reversion analysis for the determination of sequence specific effects at precise sites within a gene. |
| Databáze: | OpenAIRE |
| Externí odkaz: |
|
Full text from SpringerLink