Module 2: DNA Replication

• DNA replication involves the possible different types of DNA replication, semiconservative replication and its experimental proof, the role of topoisomerases in DNA replication, and why it's important for DNA to double to pass on equal amounts of DNA when cells divide.
• DNA replication is essential for reproduction and continued life.
• Defects in DNA replication can cause diseases like cancer.
• DNA topoisomerases solve the problem of DNA double helix unwinding caused by the separation of the two DNA strands during DNA replication.
• Topoisomerases allow efficient and accurate replication of DNA.
• In DNA replication, the two DNA strands separate and DNA is copied using the parent strand as a template.
• Complementary base pairing provides an obvious way for a double-stranded DNA molecule to replicate, this is semiconservative replication.
• Semiconservative replication results in two identical strands, each daughter strand has half the information from the parent.
• Conservative replication and dispersive replication are other possibilities, but semiconservative replication is the most common.
• In conservative replication, the two DNA strands remain together and DNA is copied using the parent strand as a template.
• In dispersive replication, the two DNA strands separate and DNA is copied using the parent strand as a template.
• DNA is plectonemic, meaning strands can’t be separated without unwinding.
• Many DNA molecules are circular, such as bacteria DNA, human mitochondria DNA, plasmids, and DNA viruses.
• The mode of action of a Type II DNA topoisomerase is to cut the G Segment, allowing the T Segment to pass through.
• 15N-DNA can be distinguished from 14N-DNA by density gradient centrifugation.
• The DNA molecule would have to rotate at 20,000 revs per minute to replicate 200,000 bp per minute.
• DNA Supercoils are produced if strands are separated, these supercoils need to be removed or prevented.
• The Meselson-Stahl experiment was carried out in 1959 to distinguish between the three modes of DNA replication and work out which occurs in E. coli bacteria.
• There are 10 bp per turn of the helix, meaning the DNA molecule would have to rotate at 20,000 revs per minute.
• The Meselson-Stahl experiment showed that in living cells, DNA is replicated by the semiconservative process.
• The mode of action of a Type I DNA topoisomerase is to separate the strands without helix unwinding during DNA replication.
• A typical bacterial chromosome is a circular DNA molecule of about 4,000,000 bp (4Mb).
• The culture medium contained 15NH4Cl - heavy ammonium chloride, causing all of the bacteria’s DNA to become labelled with 15N.
• If a bacterial chromosome is converted to a linear molecule, a bacterium can divide every 20 minutes, meaning it must replicate its genome every 20 minutes.
• Unwinding a circular molecule is a problem, maybe you make a double-stranded cut and then unwind it from the two ends, but this is linear.
• DNA polymerases are the cellular enzymes that can make DNA strands from nucleotides.
• Mutations in DNA polymerases can cause colorectal cancer.
• DNA replication can only occur in one direction but the two strands at the replication fork are in opposite orientations.
• DNA replication occurs by the semiconservative method.
• DNA topoisomerases nick/cut the DNA to relieve the stress of DNA unwinding during replication.
• Helicases break the base pairs during replication.
• The replication fork progresses along the DNA molecule during replication.
• Template-dependent DNA synthesis is the process by which new strands of DNA are synthesized at the replication fork.
• The template for template-dependent DNA synthesis is DNA, so this is DNA-dependent DNA synthesis.
• DNA-dependent DNA synthesis is carried out by an enzyme called a DNA-dependent DNA polymerase.
• DNA synthesis is always in the 5´→3´ direction, which requires a primer to initiate synthesis of the new strand.
• Without a primer, there is no DNA synthesis.
• Most DNA polymerases can also degrade DNA, this is called the exonuclease activity.
• There are two possible types of exonuclease activity: 3´→5´exonuclease activity, which allows the polymerase to remove nucleotides it has just inserted, and 5´→3´exonuclease activity, which allows the polymerase to remove DNA already attached to the template.
• DNA pol III makes the new strand once the primer has been made.