ORS12, a mammalian autonomously replicating DNA sequence, is present at the centromere of CV-1 cell chromosomes.
ors12, an 812-bp-long sequence, previously isolated by extrusion of nascent DNA from replication bubbles active at the onset of S phase (G. Kaufmann, M. Zannis-Hadzopoulus, and R. G. Martin Mol. Cell. Biol. 5, 721-727, 1985), has been shown to function as an origin of DNA replication in autonomously replicating plasmids (L. Frappier and M. Zannis-Hadjopoulos Proc. Natl. Acad. Sci. USA 84, 6668-6672, 1987) and in a cell-free system (C. E. Pearson, L. Frappier, and M. Zannis-Hadzopoulos Biochim. Biophys. Acta 1090, 156-166, 1991). A portion of ors12 (nucleotides 1-168) consists of the highly reiterated alpha-satellite sequence (B. S. Rao et al. Gene 87, 233-242, 1990). We have estimated the copy number of the non-alpha-satellite portion of ors12 in CV-1 cells to be < 9 copies per haploid genome and have used it as a probe to generate a genomic map of ors12 on CV-1 DNA. In situ hybridization of CV-1 metaphase chromosomes, using a biotinylated probe of the entire ors12 sequence, positively identified the centromeres of all chromosomes. However, when the non-alpha-satellite portion of ors12 was used as a probe, it positively identified the centromeric region of only six chromosomes, namely, B4, C11, D14, D24, E25, and E27, as well as that of a marker chromosome. The results suggest that ors12 represents a centromeric putative replication origin that is present on a subset of CV-1 chromosomes and is activated at the onset of S phase.
Mah
D C
DC
McGill Cancer Centre, Department of Medicine, McGill University, Montréal Québec, Canada.
Shihab-el-Deen
A
A
Price
G B
GB
Zannis-Hadjopoulos
M
M
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002503:Centromere / Q000737:chemistry*; D002522:Chlorocebus aethiops; D002874:Chromosome Mapping; D004247:DNA / Q000032:analysis*; D015342:DNA Probes; D004261:DNA Replication; D004276:DNA, Satellite / Q000032:analysis*; D006367:HeLa Cells; D006801:Humans; D017403:In Situ Hybridization; D008969:Molecular Sequence Data
D015342:DNA Probes; D004276:DNA, Satellite; D004247:DNA