Meselson and Stahl used DNA labeled with heavy and light nitrogen isotopes to show that DNA replicates semiconservatively. The method of analysis was cesium chloride equilibrium density gradient centrifugation. When double-stranded DNA is heated to 100c, the two strands separate (denaturation). As solution cools, the complementary strands bind and reform a double helix (renaturation). If a mixture of 15N-containing and 14N-containing DNA is heated to 100c and then cooled slowly before centrifuging, two bands are seen in the same positions as Meselson and Stahl observed, but a new third band is seen as well.

A- Explain the existence of the three bands in the gradient prior to centrifugation.

B- What does this third band represent at a molecular level?

C- Where does this band appear relative to the two bands observed by Meselson and Stahl?

D- Why does this band appear prior to centrifugation but not after?