Which experimental methods can be used to study DNA replication forks?

• A. DNA fiber assays (molecular combing), 2D gel electrophoresis of replication intermediates, electron microscopy of replication structures, and BrdU incorporation assays.
• B. Western blotting, Sanger sequencing, CRISPR knockouts, and flow cytometry.
• C. Northern blot, RT-qPCR, microarray, RNA-seq.
• D. Mass spectrometry, NMR, cryo-EM, X-ray crystallography.

Answer: (A)

When studying DNA replication forks, you need tools that reveal the fork itself and how DNA synthesis proceeds. DNA fiber assays (molecular combing) stretch individual DNA molecules and label newly made DNA, letting you see fork progression, speed, stalling, and restart at the single-molecule level. Two-dimensional gel electrophoresis of replication intermediates separates DNA forms produced during replication, such as forked and bubbled structures, so you can detect where forks progress, pause, or originate. Electron microscopy of replication structures provides direct images of fork geometry and related intermediates, giving a visual readout of fork architecture. BrdU incorporation assays label newly synthesized DNA with a thymidine analog, which can be detected to measure DNA replication activity and timing within cells.

Other approaches listed focus on proteins, RNA, or general structural study of biomolecules rather than directly showing fork structure or fork dynamics, so they don’t provide the same fork-specific insights.