Cellular Biology>Cell Cycle Mitosis Meiosis Practice Test

Researchers pulse-label asynchronously dividing mammalian fibroblasts with the thymidine analog BrdU for 30 minutes and then fix the cells. Immunofluorescence reveals punctate nuclear foci corresponding to replication factories, and PCNA localizes to these sites alongside DNA polymerase delta. Western blots show activation of S-phase cyclin-dependent kinases, while chromatin fractionation indicates robust association of the MCM helicase complex. Chromosomes are not visibly condensed, and the nuclear envelope remains intact; there is no evidence of spindle formation or kinetochore-microtubule attachments. Cohesin is detected loading behind replication forks, establishing cohesion between newly synthesized sister chromatids as DNA synthesis progresses. Flow cytometry demonstrates a shift in cellular DNA content from 2N toward intermediate values during the pulse, with aphidicolin treatment in parallel cultures blocking incorporation of the analog. After a brief chase, labeled DNA relocates consistent with known replication timing domains. Investigators conclude that the observed incorporation of a nucleotide analog and establishment of sister chromatid cohesion occur during the phase when DNA is actively synthesized.

A team investigates homologous recombination in primary spermatocytes. Using chromosome spreads, they observe pairing of homologous chromosomes along their lengths and assembly of the synaptonemal complex. Spo11-dependent double-strand breaks are detected and subsequently processed to form Holliday junction intermediates that are resolved into crossovers. Immunostaining reveals chiasmata, which physically link homologs and ensure proper alignment later in division. The nuclear envelope remains intact during these events, microtubules have not yet attached to kinetochores, and no poleward movement of chromosomes is observed. Later, when the synaptonemal complex disassembles, chiasmata persist as the sole connections between homologs. The researchers emphasize that the exchange of genetic material occurs before homologous pairs align on the spindle equator and that this exchange promotes both genetic diversity and proper segregation in the first meiotic division.