Basics of DNA Replication


Related Posts

Illustration shows the conservative, semi-conservative, and dispersive models of D N A synthesis. In the conservative model, the D N A is replicated and both newly synthesized strands are paired together. In the semi-conservative model, each newly synthesized strand pairs with a parent strand. In the dispersive model, newly synthesized D N A is interspersed with parent D N A within both D N A strands.
The three suggested models of DNA replication. Gray indicates the original DNA strands, and blue indicates newly synthesized DNA. Source: OpenStax Biology 2e

OpenStax Biology 2e

The elucidation of the structure of the double helix provided a hint as to how DNA divides and makes copies of itself. In their 1953 paper, Watson and Crick penned an incredible understatement: “It has not escaped our notice that the specific pairing we have postulated immediately suggests a possible copying mechanism for the genetic material.” With specific base pairs, the sequence of one DNA strand can be predicted from its complement. The double-helix model suggests that the two strands of the double helix separate during replication, and each strand serves as a template from which the new complementary strand is copied. What was not clear was how the replication took place. There were three models suggested: conservative, semi-conservative, and dispersive.

In conservative replication, the parental DNA remains together, and the newly formed daughter strands are together. The semi-conservative method suggests that each of the two parental DNA strands acts as a template for new DNA to be synthesized; after replication, each double-stranded DNA includes one parental or “old” strand and one “new” strand. In the dispersive model, both copies of DNA have double-stranded segments of parental DNA and newly synthesized DNA interspersed.

Meselson and Stahl were interested in understanding how DNA replicates. They grew E. coli for several generations in a medium containing a “heavy” isotope of nitrogen (15N), which gets incorporated into nitrogenous bases, and eventually into the DNA.

Illustration shows an experiment in which E coli was grown initially in media containing superscript 15 baseline upper case N  nucleotides. When the D N A was extracted and run in an ultracentrifuge, a band of D N A appeared low in the tube. The culture was next placed in the superscirpt 14 baseline upper case N medium. After one generation, all of the D N A appeared in the middle of the tube, indicating that the D N A was a mixture of half superscript 14 baseline upper N and half superscript 15 baseline upper N, D N A. After two generations, half of the D N A appeared in the middle of the tube, and half appeared higher up, indicating that half the D N A contained 50% superscript 15 baseline upper N, and half contained superscript 14 baseline upper N only. In subsequent generations, more and more of the D N A appeared in the upper, superscript 14 baseline upper N band.
Meselson and Stahl experimented with E. coli grown first in heavy nitrogen (15N) then in 14N. DNA grown in 15N (red band) is heavier than DNA grown in 14N (orange band), and sediments to a lower level in cesium chloride solution in an ultracentrifuge. When DNA grown in 15N is switched to media containing 14N, after one round of cell division the DNA sediments halfway between the 15N and 14N levels, indicating that it now contains fifty percent 14N. In subsequent cell divisions, an increasing amount of DNA contains 14N only. These data support the semi-conservative replication model. (credit: modification of work by Mariana Ruiz Villareal)

The E. coli culture was then placed into medium containing 14N and allowed to grow for several generations. After each of the first few generations, the cells were harvested and the DNA was isolated, then centrifuged at high speeds in an ultracentrifuge. During the centrifugation, the DNA was loaded into a gradient (typically a solution of salt such as cesium chloride or sucrose) and spun at high speeds of 50,000 to 60,000 rpm. Under these circumstances, the DNA will form a band according to its buoyant density: the density within the gradient at which it floats. DNA grown in 15N will form a band at a higher density position (i.e., farther down the centrifuge tube) than that grown in 14N. Meselson and Stahl noted that after one generation of growth in 14N after they had been shifted from 15N, the single band observed was intermediate in position in between DNA of cells grown exclusively in 15N and 14N. This suggested either a semi-conservative or dispersive mode of replication. The DNA harvested from cells grown for two generations in 14N formed two bands: one DNA band was at the intermediate position between 15N and 14N, and the other corresponded to the band of 14N DNA. These results could only be explained if DNA replicates in a semi-conservative manner. And for this reason, therefore, the other two models were ruled out.

During DNA replication, each of the two strands that make up the double helix serves as a template from which new strands are copied. The new strands will be complementary to the parental or “old” strands. When two daughter DNA copies are formed, they have the same sequence and are divided equally into the two daughter cells.

How is DNA Replicated?

Replication occurs in three major steps: the opening of the double helix and separation of the DNA strands, the priming of the template strand, and the assembly of the new DNA segment. During separation, the two strands of the DNA double helix uncoil at a specific location called the origin. Several enzymes and proteins then work together to prepare, or prime, the strands for duplication. Finally, a special enzyme called DNA polymerase organizes the assembly of the new DNA strands. The following description of this three-stage process applies generally to all cells, but specific variations within the process may occur depending on organism and cell type.


Clark, M., Douglas, M., Choi, J. Biology 2e. Houston, Texas: OpenStax. Access for free at:

Research Abstract: Origins of DNA replication

In all kingdoms of life, DNA is used to encode hereditary information. Propagation of the genetic material between generations requires timely and accurate duplication of DNA by semiconservative replication prior to cell division to ensure each daughter cell receives the full complement of chromosomes. DNA synthesis of daughter strands starts at discrete sites, termed replication origins, and proceeds in a bidirectional manner until all genomic DNA is replicated. Despite the fundamental nature of these events, organisms have evolved surprisingly divergent strategies that control replication onset. Here, we discuss commonalities and differences in replication origin organization and recognition in the three domains of life.

Cell Size and the Initiation of DNA Replication in Bacteria

In eukaryotes, DNA replication is coupled to the cell cycle through the actions of cyclin-dependent kinases and associated factors. In bacteria, the prevailing view, based primarily from work in Escherichia coli, is that growth-dependent accumulation of the highly conserved initiator, DnaA, triggers initiation. However, the timing of initiation is unchanged in Bacillus subtilis mutants that are ∼30% smaller than wild-type cells, indicating that achievement of a particular cell size is not obligatory for initiation. Prompted by this finding, we re-examined the link between cell size and initiation in both E. coli and B. subtilis. Although changes in DNA replication have been shown to alter both E. coli and B. subtilis cell size, the converse (the effect of cell size on DNA replication) has not been explored. Here, we report that the mechanisms responsible for coordinating DNA replication with cell size vary between these two model organisms. In contrast to B. subtilis, small E. coli mutants delayed replication initiation until they achieved the size at which wild-type cells initiate. Modest increases in DnaA alleviated the delay, supporting the view that growth-dependent accumulation of DnaA is the trigger for replication initiation in E. coli. Significantly, although small E. coli and B. subtilis cells both maintained wild-type concentration of DnaA, only the E. coli mutants failed to initiate on time. Thus, rather than the concentration, the total amount of DnaA appears to be more important for initiation timing in E. coli. The difference in behavior of the two bacteria appears to lie in the mechanisms that control the activity of DnaA.

An Increase in Mitochondrial DNA Promotes Nuclear DNA Replication in Yeast

Coordination between cellular metabolism and DNA replication determines when cells initiate division. It has been assumed that metabolism only plays a permissive role in cell division. While blocking metabolism arrests cell division, it is not known whether an up-regulation of metabolic reactions accelerates cell cycle transitions. Here, we show that increasing the amount of mitochondrial DNA accelerates overall cell proliferation and promotes nuclear DNA replication, in a nutrient-dependent manner. The Sir2p NAD+-dependent de-acetylase antagonizes this mitochondrial role. We found that cells with increased mitochondrial DNA have reduced Sir2p levels bound at origins of DNA replication in the nucleus, accompanied with increased levels of K9, K14-acetylated histone H3 at those origins. Our results demonstrate an active role of mitochondrial processes in the control of cell division. They also suggest that cellular metabolism may impact on chromatin modifications to regulate the activity of origins of DNA replication.

DNA Replication Control Is Linked to Genomic Positioning of Control Regions in Escherichia coli

Chromosome replication in Escherichia coli is in part controlled by three non-coding genomic sequences, DARS1, DARS2, and datA that modulate the activity of the initiator protein DnaA. The relative distance from oriC to the non-coding regions are conserved among E. coli species, despite large variations in genome size. Here we use a combination of i) site directed translocation of each region to new positions on the bacterial chromosome and ii) random transposon mediated translocation followed by culture evolution, to show genetic evidence for the importance of position. Here we provide evidence that the genomic locations of these regulatory sequences are important for cell cycle control and bacterial fitness. In addition, our work shows that the functionally redundant DARS1 and DARS2 regions play different roles in replication control. DARS1 is mainly involved in maintaining the origin concentration, whether DARS2 is also involved in maintaining single cell synchrony.

Multiple Regulatory Systems Coordinate DNA Replication with Cell Growth in Bacillus subtilis

In many bacteria the rate of DNA replication is linked with cellular physiology to ensure that genome duplication is coordinated with growth. Nutrient-mediated growth rate control of DNA replication initiation has been appreciated for decades, however the mechanism(s) that connects these cell cycle activities has eluded understanding. In order to help address this fundamental question we have investigated regulation of DNA replication in the model organism Bacillus subtilis. Contrary to the prevailing view we find that changes in DnaA protein level are not sufficient to account for nutrient-mediated growth rate control of DNA replication initiation, although this regulation does require both DnaA and the endogenous replication origin. We go on to report connections between DNA replication and several essential cellular activities required for rapid bacterial growth, including respiration, central carbon metabolism, fatty acid synthesis, phospholipid synthesis, and protein synthesis. Unexpectedly, the results indicate that multiple regulatory systems are involved in coordinating DNA replication with cell physiology, with some of the regulatory systems targeting oriC while others act in a oriC-independent manner. We propose that distinct regulatory systems are utilized to control DNA replication in response to diverse physiological and chemical changes.


Keywords: dna replication steps, dna replication summary, dna polymerase, why is dna replication important, dna replication diagram, where does dna replication occur, replication fork, dna replication in prokaryotes

Leave a Reply

Your email address will not be published.