7.2: Semi-Conservative DNA Replication - Biology

7.2: Semi-Conservative DNA Replication - Biology

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DNA replication is similar to transcription in its most general idea: a polymerase enzyme reads a strand of DNA one nucleotide at a time, it takes a random nucleotide from the nucleoplasm, and if it is complementary to the nucleotide in the DNA, the polymerase adds it to the new strand it is creating. Of course, there are significant differences between replication and transcription too, not the least of which is that both strands of DNA are being read simultaneously in order to create two new complementary strands that will eventually result in a complete and nearly perfect copy of an entire organismal genome.

One of the most important concepts of DNA replication is that it is a semi-conservative process (Figure (PageIndex{7})). This means that every double helix in the new generation of an organism consists of one complete “old” strand and one complete “new” strand wrapped around each other. This is in contrast to the two other possible models of DNA replication, the conservative model, and the dispersive model. A conservative mechanism of replication proposes that the old DNA is used as a template only and is not incorporated into the new double-helix. Thus the new cell has one completely new double-helix and one completely old double-helix. The dispersive model of replication posits a final product in which each double helix of DNA is a mixture of fragments of old and new DNA. In light of current knowledge, it is difficult to imagine a dispersive mechanism, but at the time, there were no mechanistic models at all. The Meselson-Stahl experiments (1958) clearly demonstrated that the mechanism must be semi-conservative, and this was confirmed once the key enzymes were discovered and their mechanisms elucidated.

In the Meselson-Stahl experiments, E. coli were first incubated with 15N, a heavy isotope of nitrogen. Although it is only a difference in mass of one neutron per atom, there is a great enough difference in mass between heavy nitrogen-containing DNA (in the purine and pyrimidine bases) and light/normal nitrogen-containing DNA that they can be separated from one another by ultracentrifugation through a CsCl concentration gradient (Figure (PageIndex{7})).

Over 14 generations, this led to a population of E. coli that had heavy nitrogen incorporated into all of the DNA (shown in blue). Then, the bacteria are grown for one or two divisions in “light” nitrogen, 14N. When the DNA from the bacterial populations was examined by centrifugation, it was found that instead of light DNA and heavy DNA, as would be expected if DNA replications was conservative, there was a single band in and intermediate position on the gradient. This supports a semi-conservative model in which each strand of original DNA not only acts as a template for making new DNA, it is itself incorporated into the new double-helix.

3.4 & 7.2 DNA Replication

  • Origins of Replication:site in which replication starts
      • Prokaryotic cells have one origin, and eukaryoic cells have many.
        • nucleoside triphosphate hydrogen, bonds to complementary base in DNA molecule
        • hydrolysis of 2 phosphate molecules = convert nucleoside triphosphate to nucleotide
        • Only added at 3′ end of nucleotide = DNA strands are organized in anti-paralllel way
        • Nucleoside/ Nucleoside monophosphate: sugar and a base
        • Okazaki fragments: lagging strands
            • enzyme DNA ligase: help join fragments together
              • catalyzed by enzyme DNA polymerase III
              • needs a short sequence (primer) to start process *primer: made of RNA and primase enzyme
              • The DNA polymerase I = enzyme that digests away RNA primer + replace with DNA

              The Meselson-Stahl Experiment (p.63)

              Data in Figure 14 suggests that replication is semi-conservative because a peak is not present at 1.710 in N-15. Because the DNA molecule built with only N0-14 did not have a peak, the replication can be stated to not be conservative. The figure is not dispersive because the second generation consists of 2 peaks instead of one. Also, if it is dispersive, the all the molecules would have the same ratio of N-14 to N-15. no matter the time/generation (last figure from figure 14).

              Meselson-Stahl Experiment (p.64)

              (1) The sample that was pulsed for 10 seconds only consists of one high peak rather than the sample pulsed for 30 seconds, which had 2 peaks throughout. Also, the 3o second sample exhibited a higher radioactivity to the distance from the top, compared to the sample pulsed for 10 seconds.

              (2) The sample pulsed for 30 seconds provides evidence for the presence of both a leading strand and many lagging strands because of the two apparent peaks in radioactivity. The high peak indicates the lagging strands for its steep increase and the lower peak shows the presence of leading strands due to the longer fragments.

              (3) The sample pulsed for 60 seconds provides evidence for the activity of DNA ligase because of the number of molecules. The small molecules decreases and larger molecules would increase because the DNA ligase merge the lagging strands together.

              7.2 DNA Replication

              This reviews the Core content before going on to the AHL stuff.

              DNA Replication animations:

              More animations from North Harris College and from LearnersTV.

              Revision materials:

              Here is the top-rated video on the subject on YouTube:

              Key terms: free DNA nucleotide, 5′, 3′, replicase, helicase, RNA primase, DNA polymerase, DNA ligase, Okazaki, deoxynucleoside triphosphate,

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              Semi Conservative Theory Of Dna Replication

              1958 also demonstrated the semi conservative mode of replication in dna and chromosomes in root tip cells of vicia faba. Semiconservative replication describes the mechanism of dna replication in all known cells.

              Semi Conservative Dna Replication Youtube

              One of the most important concepts of dna replication is that it is a semi conservative process figure 7 2.

              Semi conservative theory of dna replication. Watson and crick figured that this model would result in two new double strands of dna each one with one strand of parent or template dna and one strand of daughter or newly synthesized dna. During dna replication the double stranded dna helix unwinds with the help of the enzyme helicase and the strands are separated. This is called semiconservative replication.

              Semi conservative replication posits the creation of hybrid old new double helices. This results in the formation of two identical copies of the original double stranded molecule. This results in two dna molecules with one original strand and one new strand.

              Semi conservative replication was shown to be the method of replication by meselson and stahl in 1958. Dna replication occurs on multiple origins of replication along the dna template strand. As the dna double helix is unwound by helicase replication occurs separately on each template strand in antiparallel directions.

              Dna replication is semiconservative meaning that each strand in the dna double helix acts as a template for the synthesis of a new complementary strand. They used coli a bacteria and two nitrogen isotopes a heavy form 15 n and the normal form 14 n to demonstrate how the density of dna changes over generations as the 15 n isotope was replaced with the 14 n isotope. After the incorporation of radioactive thymidine 3 h the root tips were transferred to unlabelled medium containing colchicine.

              Dispersive replication proposed molecules composed of randomized fragments of double old and double new dna. In this model the two strands of dna unwind from each other and each acts as a template for synthesis of a new complementary strand. Semiconservative replication during dna replication a double stranded dna molecule separate and each strand is used as a template for the synthesis of a new strand.

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              7.2 – DNA Replication

              7.2 – DNA Replication

              7.2.1 – State that DNA replication occurs in a 5′ → 3′ direction

              The 3’ end of the nucleotide is a free -OH group. The 5’ end of the free DNA nucleotide is added to the 3’ end of the chain of nucleotides that is already synthesised.

              7.2.2 – Explain the process of DNA replication in prokaryotes, including the role of enzymes (helicase, DNA polymerase, RNA primase and DNA ligase), Okazaki fragments and deoxynucleoside triphosphates

              In prokaryotes, the initiation spot on the DNA is called the ori, or origin point. Replications finishes at the ter spot, or termination point.

              This binds to the double helix to stimulate the separation of the strands. The hydrogen bonds between base pairs break to form replication forks. The helicase is located at these replication forks.

              Polymerase III replicates DNA in a 5’ to 3’ direction along the leading strand. It starts at the RNA primer, adding nucleotides using complementary base pairing and moving in the direction of the replication fork. On the other hand, along the lagging strand, polymerase III moves away from the replication fork. This results in the formation of Okazaki fragments. Polymerase I replaces the RNA primers with DNA. However, there is still a gap where two nucleotides have not been connected.

              It is possible that errors may occur during replication, but the polymerase has mechanisms of back-checking for mutations.

              These are short strands of DNA that are formed on the lagging strand. Each one is initiated at the replication fork, and is later joined to form one continuous length by DNA ligase. The leading strand is replicated in one continuous length

              For replication to occur, a free 3’ hydroxyl group is required. Primase synthesises at the initiation sites.

              Gaps a made in the DNA from where the primer is removed. Ligase closes the gap by forming a covalent bond between the phosphate groups and the neighbouring fragments are joined.

              The free nucleotides have three phosphates. During polymerisation, the condensation reaction, two are removed so that only one remains to form the backbone

              7.2.3 – State that DNA replication is initiated at many points in eukaryotic chromosomes

              Prokaryotic DNA is replicated in a continuous loop. On the other hand, eukaryotic DNA is replicated at multiple points to speed up the reaction. The DNA is unwound at multiple points along the helix into bubbles that expand, allowing replication to continue in both direction. The bubbles eventually fuse.