EM analysis was performed according to the standard protocol (9 (link)). For DNA extraction, cells were lysed in lysis buffer and digested at 50°C in the presence of proteinase K for 2 hours. The DNA was purified using chloroform/isoamyl alcohol, precipitated in isopropanol, given 70% ethanol wash, and resuspended in elution buffer. Isolated genomic DNA was digested with Pvu II high-fidelity restriction enzyme for 4 to 5 hours. After digestion, the DNA solution was transferred to a Microcon DNA fast flow centrifugal filter. The filter was washed with tris-EDTA (TE) buffer after spinning for 7 min. The benzyldimethylalkylammonium chloride method was used to spread the DNA on the water surface and then loaded on carbon-coated nickel grids, and last, DNA was coated with platinum using high-vacuum evaporator MED 010 (Bal-Tec). Microscopy was performed with a transmission electron microscope FEI Talos, with 4K by 4K complementary metal-oxide semiconductor camera. For each experimental condition, at least 200 replication fork intermediates were analyzed from three independent experiments, and MAPS software (Thermo Fisher Scientific) was used to analyze the images.