DNA Replication – the Replication Fork and the Meselson–Stahl Experiment
BiologyGeneticsAges 17–18
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Sign in to playAn animated replication fork: helicase unwinds the DNA, primase lays RNA primers, DNA polymerase III extends new strands 5′→3′ – continuously on the leading strand and as Okazaki fragments on the lagging strand – then DNA polymerase I replaces the primers and ligase joins the fragments. Run, pause or step through events, change the Okazaki fragment length and count primers and nicks. A second screen recreates the Meselson–Stahl experiment: E. coli grown in ¹⁵N are moved to ¹⁴N, and the DNA bands after CsCl density-gradient centrifugation in generations 0–4 are compared with the predictions of the conservative, semi-conservative and dispersive hypotheses.
Lesson: DNA replication: the replication fork, roles of the enzymes, leading and lagging strands; semi-conservative replication (Meselson–Stahl experiment)
What it shows
DNA polymerase can only add nucleotides to a free 3′-OH, so every new strand grows 5′→3′ and needs a short RNA primer to start. At a replication fork, helicase separates the strands. On the leading strand, DNA polymerase III copies continuously towards the fork; on the lagging strand it works away from the fork in Okazaki fragments, each started by primase. DNA polymerase I then swaps the RNA primers for DNA and ligase seals the nicks. The Meselson–Stahl experiment, using ¹⁵N and ¹⁴N and density-gradient centrifugation, showed that replication is semi-conservative.
How to use
On Replication fork, press Play or use Step to go one event at a time; read the event box and count primers, fragments and nicks. Change Okazaki fragment and predict how many primers will be needed. On Meselson–Stahl experiment, press Next generation, compare each tube with the three predictions and use Hypothesis to show one model at a time. The table marks which hypotheses the data rule out.
Parameters you can change
- Screen Replication fork, Meselson–Stahl experiment
- Playback speed (1 = a quarter of the real E. coli rate) 0.25–3
- Okazaki fragment length 200–2000 nt
- Show enzyme names
- Generations grown in ¹⁴N at the start (Meselson–Stahl screen) 0–4
- Hypotheses shown All three hypotheses, Semi-conservative, Conservative, Dispersive
Questions to explore
- Why must the lagging strand be made in short pieces while the leading strand is continuous?
- Which hypothesis does generation 1 rule out, and why is generation 2 needed to reject the dispersive model?
- After three generations in ¹⁴N, what percentage of the DNA is hybrid?
