Molecular biology – In a cesium chloride (CsCl) gradient centrifugation experiment, DNA molecules are separated primarily based on which physical property?

Introduction:
Cesium chloride (CsCl) density-gradient centrifugation is a classic analytical and preparative technique in molecular biology. It is famous for separating nucleic acids according to their buoyant density and for enabling the historic demonstration of semi-conservative DNA replication. This question tests the core principle behind how DNA bands form in a CsCl gradient.

Given Data / Assumptions:

• Under ultracentrifugation, CsCl forms a continuous density gradient.
• DNA migrates until it reaches a position where its density equals the surrounding solution.
• Bouyant density depends on base composition (notably GC content) and isotopic labeling (e.g., 15N vs 14N).

Concept / Approach:
When spun at high speed, CsCl solutions stratify such that each position corresponds to a specific solution density. DNA does not separate by size in this setup; instead, it moves to the exact point in the gradient where net buoyancy is zero. Differences in GC content or isotopic labeling alter buoyant density slightly, allowing discrete bands for different DNA populations.

Step-by-Step Solution:

Verification / Alternative check:
In the Meselson–Stahl experiment, 15N-labeled DNA bands at a higher density than 14N DNA, proving separation is by density rather than length or charge. Mixing populations yields distinct or hybrid-density bands exactly where predicted by buoyant-density calculations.

Why Other Options Are Wrong:

• Absorption/resorption: not the driving force for banding in CsCl.
• Adhesion to the tube: irrelevant to equilibrium band formation.
• Net electric charge: electrophoresis uses charge/size; CsCl gradients use buoyant density.

Common Pitfalls:
Confusing CsCl equilibrium density gradients with sucrose rate-zonal gradients (which separate primarily by size/shape over time, not by equilibrium density).

Final Answer:
Density (buoyant density in the CsCl gradient)