DNA replication is the fundamental process by which a cell creates an exact copy of its genetic material. This duplication is necessary before a cell divides, ensuring each new daughter cell receives a complete set of instructions. Scientists sought to understand how the intricate double-helix structure of DNA could accurately unwind and duplicate itself. This presented a puzzle, with various possibilities for how the copying mechanism might unfold.
The Three Proposed Models of Replication
Before experimental evidence clarified the process, scientists considered several ways DNA might replicate.
The conservative model proposed that the original double-stranded DNA molecule remained entirely intact after replication. The parent molecule would serve as a template, then re-form, while a completely new, identical daughter DNA molecule was synthesized from scratch. This would result in one old DNA molecule and one entirely new DNA molecule after one round of replication.
The semi-conservative model proposed that the two strands of the parent DNA double helix would separate. Each strand would then act as a template for the synthesis of a new, complementary strand. This process would yield two new DNA molecules, with each molecule consisting of one original “parental” strand and one newly synthesized “daughter” strand.
The dispersive model suggested that the parent DNA molecule would break into multiple segments during replication. The new DNA molecules would then be formed as a patchwork, containing interspersed sections of both old (parental) and new (synthesized) DNA on each strand. Each daughter DNA molecule would be a mosaic of original and new material.
The Meselson-Stahl Experiment
In 1958, Matthew Meselson and Franklin Stahl devised an experiment to determine which of these models accurately described DNA replication. They utilized Escherichia coli bacteria and distinct isotopes of nitrogen to label the DNA. Nitrogen is a component of DNA’s bases, making it suitable for labeling.
The experiment began by growing E. coli for several generations in a medium containing a “heavy” isotope of nitrogen, ¹⁵N. This heavy nitrogen was incorporated into the bacteria’s DNA, making it denser than normal DNA. After ensuring all bacterial DNA contained ¹⁵N, the bacteria were transferred to a new medium containing the “light” isotope of nitrogen, ¹⁴N.
As the bacteria grew and divided in the ¹⁴N medium, samples were collected after one and two rounds of DNA replication. The DNA was extracted from these samples and analyzed using density-gradient centrifugation. This technique separates molecules based on their density by spinning DNA samples at high speeds in a cesium chloride solution.
Experimental Results and Conclusions
After one round of replication in the ¹⁴N medium, Meselson and Stahl observed that the extracted DNA formed a single band during density-gradient centrifugation. This band was positioned at an intermediate density, falling between the expected densities of pure ¹⁵N-labeled DNA and pure ¹⁴N-labeled DNA. This result immediately disproved the conservative model of DNA replication. If DNA replication were conservative, the first generation would have produced two distinct bands: one heavy band representing the original ¹⁵N parental DNA and one light band representing the entirely new ¹⁴N daughter DNA.
The single intermediate band was consistent with both the semi-conservative and dispersive models. To differentiate, the scientists allowed the bacteria to undergo a second round of replication in the ¹⁴N medium. The DNA extracted from this second generation yielded two distinct bands after centrifugation. One band remained at the intermediate density, identical to the band observed in the first generation. The other band was lighter, corresponding to the density of DNA composed entirely of ¹⁴N.
These second-generation results disproved the dispersive model. The dispersive model would have predicted that all DNA molecules would continue to be a mixture of old and new DNA, resulting in a single band that gradually became lighter with each successive generation, not two separate bands. The appearance of both an intermediate band and a purely light band matched the predictions of the semi-conservative model. The experiment confirmed that DNA replicates by a semi-conservative mechanism, where each new DNA molecule consists of one original strand and one newly synthesized strand.