Abstract
We have used density shift analysis to monitor the autonomous replicating sequence (ARS) activity of plasmids containing various DNA fragments from the 5′-flanking region of the human c-myc gene. The ARS activity of certain of these plasmids implied that structures in the c-myc DNA could be recognized for the initiation of replication in the absence of chromosomal integration. The plasmid pNeo.Myc-2.4 contains 2.4 kb of c-myc 5′-flanking DNA, and replicated semiconservatively as a circular extrachromosomal element. Deletion derivatives of pNeo.Myc-2.4 containing either of two nonoverlapping regions of c-myc DNA semiconservatively incorporated bromodeoxyuridine into discrete populations of heavy-light supercoiled molecules to roughly the same extent as the chromosomal DNA in the same cultures. Some constructs displayed lower ARS activity, implying that distinct cis-acting sequences in the c-myc 5′-flanking DNA may independently affect DNA replication. The ARS activity of two separate c-myc sequences suggests that replication initiation signals are redundant in the c-myc origin. The smallest c-myc insert that displayed substantial ARS activity was 930 bp long and contained three 10/11 matches to the yeast ARS consensus and several additional features found in eukaryotic replication origins.
Get full access to this article
View all access options for this article.