Bi-directional replication and random termination

Two-dimensional (2D) agarose gel electrophoresis was used to study terminat ion of DNA replication in a shuttle vector, YRp7', when it replicated in Es cherichia coli, Saccharomyces cerevisiae and Xenopus egg extracts. In E,col i, the 2D gel patterns obtained were consistent with uni-directional replic ation initiated at a specific site, the ColE1 origin, In consequence, termi nation also occurred precisely at the ColE1 origin, In Xenopus egg extracts , the particular shape of the bubble are as well as the triangular smear de tected to the left of the simple-Y pattern indicated random initiation and termination. In S. cerevisiae, initiation occurred at the ARS1 origin and r eplication proceeded in a bi-directional manner, However, termination did n ot always occur at a specific site 180 degrees across from the origin, but almost all along the south hemisphere of the plasmid, Inversion, deletion o r replacement of DNA sequences located throughout this hemisphere did not e liminate random termination. Analysis of the replication intermediates of a nother yeast plasmid bearing a different origin, ARS305, also exhibited ran dom termination. We propose that the random termination events observed in S,cerevisiae could be due to an asynchronous departure of both forks from t he bidirectional origin in addition to differences in the rate of fork prog ression. These observations could be extended to all bi-directional origins .