Deoxyribonucleic acid, or DNA, holds the hereditary instructions for the development, functioning, growth, and reproduction of all known organisms. For a cell to divide and pass on its genetic blueprint to two daughter cells, the entire molecule must be copied with extraordinary accuracy. DNA replication ensures that every new cell receives a complete and faithful set of these instructions. This molecular duplication allows genetic information to be transferred from one generation of cells to the next without loss or error.
The Semi-Conservative Nature of DNA Replication
The key to understanding the origin of each strand in the replicated DNA lies in a concept known as semi-conservative replication. This model dictates that when a double-stranded DNA molecule is copied, the resulting two new DNA molecules are not entirely new constructions. Instead, each new double helix conserves half of the original material.
Each replicated DNA molecule is therefore a hybrid, consisting of one strand from the original, parental DNA and one newly synthesized strand. The term “semi-conservative” describes this mechanism because half of the original molecule is conserved in each of the two copies. The two strands of the parent DNA separate and each acts as a template for the synthesis of a complementary partner strand, ensuring the genetic information is duplicated precisely.
Initiating Replication: Unwinding the Double Helix
The first step in copying the DNA molecule is separating the two intertwined parental strands, a process that must begin at specific starting points. These designated start locations are called origins of replication, which are scattered throughout the long DNA strands in complex organisms. In regions rich in adenine (A) and thymine (T) base pairs, the double helix is easier to pull apart.
At an origin, a specialized enzyme called helicase is recruited to begin the unwinding process. Helicase moves along the double helix, breaking the hydrogen bonds that hold the complementary base pairs together. This action separates the two parental strands, creating a Y-shaped structure known as the replication fork.
The unwinding process proceeds in two opposite directions from the origin, creating a replication bubble. Proteins bind to the separated single strands to prevent them from snapping back together before the copying process can be completed.
Building the New Strands: Leading and Lagging Synthesis
With the parental strands separated, the construction of the new complementary strands begins, governed by the enzyme DNA polymerase. This enzyme can only add new nucleotides in one specific chemical direction, from the 5′ end to the 3′ end of the newly growing strand. This constraint is complicated by the fact that the two parental template strands run in opposite orientations, meaning they are antiparallel.
To solve this directional challenge, two distinct methods of synthesis are employed at the replication fork. Before DNA polymerase can start, a small segment of RNA, known as a primer, must be synthesized by the enzyme primase to provide a starting point. Once the primer is in place, DNA polymerase takes over.
Leading Strand Synthesis
The strand oriented to allow DNA polymerase to move continuously in the same direction as the advancing replication fork is called the leading strand. On this template strand, which runs in the 3′ to 5′ direction, the new strand is synthesized smoothly and without interruption, requiring only a single primer to begin the process.
Lagging Strand Synthesis
The other parental strand, oriented in the 5′ to 3′ direction, forces the synthesis to occur in the direction opposite to the fork’s movement, creating the lagging strand. Since DNA polymerase must still synthesize in the 5′ to 3′ direction, it must repeatedly start and stop, working backward in short segments. These short, newly synthesized fragments of DNA are called Okazaki fragments.
Each Okazaki fragment requires its own RNA primer to initiate synthesis. Once DNA polymerase finishes a fragment and runs into the next one, the RNA primers must be removed and replaced with DNA nucleotides. Finally, an enzyme called DNA ligase seals the gaps between the Okazaki fragments to create a single, continuous lagging strand.
Confirming the Model: The Meselson-Stahl Experiment
The hypothesis of semi-conservative replication was definitively proven by the landmark Meselson-Stahl experiment in 1958. Before this experiment, scientists considered two other possibilities: the conservative model, where the original DNA remained intact and a completely new molecule was made, and the dispersive model, where the original material was broken up and scattered throughout the new strands.
Matthew Meselson and Franklin Stahl used different isotopes of nitrogen, a major component of DNA, to tag the parental and newly synthesized strands. They grew bacteria in a medium containing a heavy isotope, \(^{15}N\), which incorporated into the bacterial DNA, making it denser. These bacteria were then transferred to a medium containing the lighter, more common isotope, \(^{14}N\).
After one round of replication, the extracted DNA showed a single band of intermediate density when separated by density gradient centrifugation. This result eliminated the conservative model, which would have produced two separate bands of heavy and light DNA.
After a second round of replication, the DNA separated into two bands: one intermediate and one light. This outcome was consistent only with the semi-conservative model, as the intermediate band represented the hybrid molecules (one old \(^{15}N\) strand and one new \(^{14}N\) strand), and the light band represented the molecules consisting entirely of two new \(^{14}N\) strands.