Journal
GENOMICS
Volume 84, Issue 3, Pages 475-484Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ygeno.2004.04.008
Keywords
replication origin; HPRT; functional homology; quantitative PCR
Funding
- NCI NIH HHS [CA84493] Funding Source: Medline
- NHLBI NIH HHS [HL37001] Funding Source: Medline
- NIEHS NIH HHS [ES09112] Funding Source: Medline
- NIGMS NIH HHS [GM2009] Funding Source: Medline
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A functional origin of replication was mapped to the transcriptional promoter and exon I of the hypoxanthine-guanine phosphoribosyltransferase (HPRT) gene in the mouse and human genomes. This origin was lost in mouse embryonic stem (ES) cells with a spontaneous deletion (similar to36 kb) at the 5' end of the HPRT locus. Restoration of HPRT activity by homologous recombination with human/mouse chimeric sequences reconstituted replication origin activity in two independent ES cell lines. Quantitative PCR analyses of abundance of genetic markers in size-fractionated nascent DNA indicated that initiation of DNA replication coincided with the site of insertion in the mouse genome of the 335 bp of human DNA containing the HPRT exon I and a truncated promoter. The genetic information contained in the human sequence and surrounding mouse DNA was analyzed for cis-acting elements that might contribute to selection and functional activation of a mammalian origin of DNA replication. (C) 2004 Elsevier Inc. All rights reserved.
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