Figure 4.

Analysis of ARS309 replication intermediates. Genomic DNA was digested with BglII, electrophoresed, and blotted as described by Brewer and Fangman (5). The 3.6-kb BglII fragment containing ARS309 was detected by probing with the 1.4-kb HindIII–EcoRI fragment indicated by the dashed lines in C. The 0.35-kb HpaI–EcoRI fragment containing ARS309 is denoted by the box. (A) Wild-type ARS309. The arrows point to bubble-shaped replication intermediates, indicating that ARS309 is active as an origin on the chromosome. (B) ARS309 with the essential 9 of 11 match mutated (Fig. 2G). There are no bubble-shaped replication intermediates (arrows), demonstrating that origin function is abolished in the mutant.