DNA replication

• • DNA copies itself before cell division so that each new cell has the full amount of DNA
  • The method is called semi-conservative replication because half of the strands in each new DNA molecule are from the original DNA molecule
  • This means that there's genetic continuity between generations of cells
• Stages of semi-conservative replication

  1. The enzyme DNA helicase breaks the hydrogen bonds between bases on the two polynucleotide DNA strands. This makes the helix unwind to form two single strands
  2. Each original strand acts as a template for a new strand. Complementary base pairing means that free DNA nucleotides are attracted to their complementary exposed bases on each original template strand
  3. Condensation reactions join the nucleotides of the new strand together - catalysed by the enzyme DNA polymerase. Hydrogen bonds form between bases on the original and new strands.
  4. Each new DNA molecule contains one strand from the original DNA molecule and one new strand
• The action of DNA polymerase

  • Each end of a DNA strand is slightly different in its structure
  • One end is called the 3' end and one end is called the 5' end
  • During DNA replication the active site of DNA polymerase is only complementary to the 3' end of the newly forming DNA strand - so the enzyme can only add nucleotides at the new strand at the 3' end
  • This means that the new strand is made in a 5' to 3' direction and that DNA polymerase moves down the template strand in a 3' to 5' direction
  • Because the strands in the double helix are antiparallel, the DNA polymerase working on one of the template strands move in the opposite direction to the DNA polymerase working on the other template strand
• Meselson and Stahl showed DNA is replicated using the semi-conservative method. Their experiment used two isotopes of nitrogen - heavy nitrogen 15N and light nitrogen 14N
• 1. Two samples of bacteria were grown for many generations - one in a nutrient broth containing light nitrogen, and one in a broth with heavy nitrogen. As the bacteria reproduced, they took up nitrogen from the broth to help make nucleotides for new DNA. So the nitrogen gradually became part of the bacterias DNA
  2. A sample of DNA was taken from each bacteria and spun in a centrifuge. The DNA from the heavy nitrogen bacteria settled lower down the centrifuge tube than the DNA from the light nitrogen bacteria because its heavier
  3. Then the bacteria grown in the heavy nitrogen broth were taken out and put in a broth containing only light nitrogen. The bacteria were left for one round of replication, and then another DNA sample was taken out and spun in the centrifuge
• 4. If replication was conservative, the original heavy DNA, which would still be together, would settle at the bottom and the new light DNA would settle at the top

  5. If replication was semi-conservative, the new bacterial DNA molecules would contain one strand of the old DNA containing heavy nitrogen and one strand of new DNA containing light nitrogen. So the DNA would settle out between where the light nitrogen DNA settled out and where the heavy nitrogen DNA settled out

  6. As it turned out, the DNA settled out in the middle, showing that the DNA molecules contained a mixture of heavy and light nitrogen. The bacterial DNA had replicated semi-conservatively in the light nitrogen.