Complementation of replication origin function in mouse embryonic stem cells by human DNA sequences

被引：9
作者：
Cohen, SM [1 ]
Hatada, S [1 ]
Brylawski, BP [1 ]
Smithies, O [1 ]
Kaufman, DG [1 ]
Cordeiro-Stone, M [1 ]
机构：
[1] Univ N Carolina, Sch Med, Dept Pathol & Lab Med, Chapel Hill, NC 27599 USA
来源：
GENOMICS | 2004年 / 84卷 / 03期
关键词：
replication origin; HPRT; functional homology; quantitative PCR;
D O I：
10.1016/j.ygeno.2004.04.008
中图分类号：
Q81 [生物工程学（生物技术）]; Q93 [微生物学];
学科分类号：
071005 ; 0836 ; 090102 ; 100705 ;
摘要：
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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收藏
页码：475 / 484
页数：10
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