Experiments in biological research can be messy. To begin with, biological systems are inherently highly variable, and you need to have good control. Then there come reagents that are so finicky that they need a super pampered buffer solution. Everything has to be in just condition and then there is the challenge of reproduction. That’s why, when someone outsider reads about experiments, s/he can soon be lost in the details. But there are exceptions – there is a handful of experiments that are not that complex and super easy to understand, yet they solve some deep mystery about life. One of them is the Meselson-Stahl experiment which proves that DNA replication is semiconservative (1958). The Meselson-Stahl experiment has been called the most beautiful experiment in biology for the elegant logic of its deceptively simple design.
Before jumping into the details of the experiment, let’s talk about the background a little bit. It was eve for molecular biology, elusive DNA structure-puzzled was solved pretty recently (1952). However, the details of how DNA copies itself are missing. At that time, three hypotheses were proposed for the method of replication of DNA. They are:
- In the semiconservative replication hypothesis, Watson and Crick proposed that the two strands of a DNA molecule separate during replication. Each strand then acts as a template for synthesizing a new strand.
- The conservative replication hypothesis proposed that the entire DNA molecule acted as a template for synthesizing an entirely new one. According to this model, histone proteins bind to the DNA, revolving the strand and exposing the nucleotide bases (which generally line the interior) for hydrogen bonding.
- The dispersive replication hypothesis is exemplified by a model proposed by Max Delbrück, which attempts to solve the problem of unwinding the two strands of the double helix by a mechanism that breaks the DNA backbone every ten nucleotides or so, untwists the molecule, and attaches the old strand to the end of the newly synthesized one. This would synthesize the DNA in short pieces alternating from one strand to the other.
Each of these three models makes a different prediction about the distribution of the “old” DNA in molecules formed after replication. To solve this puzzle, Meselson-Stahl set up the following experiment.
Nitrogen is a principal constituent of DNA. Escherichia coli cells were grown in a heavy isotope containing Nitrogen (15N) media for a prolonged period. Then those cells were again grown in a light isotope containing Nitrogen media (14N). As a result, a density gradient (some of the chromosomes were 15N labeled and some were 14N labeled) was introduced into the DNA of E. coli. In the next step, Meselson and Stahl separated the DNA by “density-gradient centrifugation.”
Note that centrifugation is a method of separating molecules having different densities by spinning them in solution around an axis (in a centrifuge rotor) at high speed. They use CsCl salt (Cesium Chloride) as a density concentrated solution in this density-gradient centrifugation process. Because the molecular weight of 6M CsCl (1.70 g/cm3) is approximately equal to the molecular weight of an E. coli (1.710 g/cm3). After ultracentrifugation DNA of E. coli was separated into different layers according to their molecular weight. From these layers, Meselson and Stahl isolated the DNAs.
After isolation, they observed that the density of that DNA was between 15N and 14N labeled DNA. This type of intermediate density is called hybrid density. Then they grow their observed DNA in 14N labeled growth media for the second generation. They got 50% hybrid DNA and 50% light DNA.
The result was consistent with the semiconservative replication hypothesis. Different results would have been obtained if DNA replication in E. coli were either conservative or dispersive. Thus, their result proved the semiconservative replication method of Watson and Crick. Till now, their experiment is famous as it is called the most beautiful experiment in biology.