oriGNAI3: A narrow zone of preferential replication initiation in mammalian cells identified by 2D gel and competitive PCR replicon mapping techniques

The nature of mammalian origins of DNA replication remains controversial and this is primarily because two-dimensional gel replicon mapping techniques have identified broad zones of replication initiation whereas several other techniques, such as quantitative PCR, have disclosed more discrete sites...

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Bibliographic Details
Published in:Nucleic acids research 1998-05, Vol.26 (10), p.2313-2321
Main Authors: Toledo, Franck, Baron, Bruno, Fernandez, Maria-Aparecida, Lachagès, Anne-Marie, Mayau, Véronique, Buttin, Gérard, Debatisse, Michelle
Format: Article
Language:English
Subjects:
AMP Deaminase - genetics
Animals
CHO Cells
Chromosome Mapping
Cricetinae
DNA Replication - genetics
Electrophoresis, Gel, Two-Dimensional - methods
GTP-Binding Proteins - genetics
Life Sciences
Molecular Sequence Data
Polymerase Chain Reaction - methods
Replication Origin - genetics
Replicon
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Summary:The nature of mammalian origins of DNA replication remains controversial and this is primarily because two-dimensional gel replicon mapping techniques have identified broad zones of replication initiation whereas several other techniques, such as quantitative PCR, have disclosed more discrete sites of initiation at the same chromosomal loci. In this report we analyze the replication of an amplified genomic region encompassing the 3′-end of the GNAI3 gene, the entire GNAT2 gene and the intergenic region between them in exponentially growing Chinese hamster fibroblasts. These cells express GNAI3 but not GNAT2. The replication pattern was first analyzed by two-dimensional neutral-alkaline gel electrophoresis. Surprisingly, the results revealed a small preferential zone of replication initiation, of at most 1.7 kb, located in a limited part of the GNAI3-GNAT2 intergenic region. Mapping of this initiation zone was then confirmed by quantitative PCR. The agreement between the two techniques exploited here strengthens the hypothesis that preferred sites of replication initiation do exist in mammalian genomes.
ISSN:0305-1048
1362-4962
DOI:10.1093/nar/26.10.2313