Lecture 2- DNA replication, the cell cycle and mitosis Flashcards
What is semi-conservative replication?
Each daughter cell inherits one old and one new strand. Each old strand acts as a template for the synthesis of the new strand –> identical copies ensured
How is the DNA helix opened up?
- DNA helicase unwinds and unzips the DNA double helix.
- Uses energy from ATP to break the hydrogen bonds between base pairs
How is the new strand of DNA synthesised?
DNA polymerase adds nucleotides to the 3’ end of DNA. DNA synthesis therefore occurs in the 5’ to 3’ direction. dNTP is added to the free 3’ -OH group
And which three things are needed by the enzyme involved in this reaction?
Three things needed:
1) Template strand
2) Oligonucleotide Primer- made of RNA by an RNA polymerase (DNA primase)
3) dNTPs- nucleotide with three phosphates. Hydrolysis of the phosphates releases energy to drive the reaction.
Explain how nucleoside analogs can be used as drugs
Eg. Acyclovir used for Herpes. The pentose sugar does not have a -OH on the 3’. Next dNTP cannot be added to the chain. Acts as a chain terminator so DNA synthesis stops.
Describe the process of DNA replication for the leading strand w.r.t:
Origin of replication, replication fork, leading strand, lagging strand (5pts)
1) Replication begins at origin of replication
2) DNA synthesis occurs at a site called the replication fork which moves with DNA replication.
3) Two daughter strands have opposite orientations.
4) Leading strand has 3’ nearest replication fork. This is formed continuously because DNA synthesised in 5’ –> 3’ direction.
5) RNA primer synthesised by DNA primase on 3’ end of template strand at replication origin and DNA polymerase binds and forms leading strand continuously.
5) Lagging strand has 3’ furthest away from replication fork. Formed discontinuously as Okazaki fragments.
Describe how DNA is synthesised on the lagging strand w.r.t:
lagging strand, Okazaki fragments, primer, DNA primase, template strand, Ribonulease, repair DNA polymerase, DNA ligase (6pts)
1) Lagging strand has 3’ furthest away from replication fork. Formed discontinuously as Okazaki fragments.
2) RNA primer, synthesised by DNA primase, attaches to template strand.
3) DNA polymerase adds nucleotides to 3’ end of primer and continues to synthesise Okazaki fragment until it reaches the end of the next fragment.
4) Old RNA primer removed by ribonuclease using 5’ to 3’ exonuclease activity
5) Repair DNA polymerase replaces RNA with DNA
6) DNA ligase- joins the Okazaki fragments together.
7) The lagging strand forms a loop so that both strands can be synthesised simultaneously.
What is the purpose of the single strand DNA binding protein?
To prevent single stranded DNA from locally folding
What is the function of the sliding clamp?
To make sure DNA polymerase is in the right place
Explain how accuracy of DNA synthesis is maintained?
- DNA polymerase has proof-reading mechanism.
- Before new nucleotide is added, the previous nucleotide is checked for correct base-pairing.
- Incorrect bases are removed by 3’ to 5’ exonuclease activity of DNA polymerase
Describe how E. coli chromosome replication occurs.
1) Replication starts at OriC
2) Two replication forks
3) Get Bi-directional replication
Describe the various stages of the cell cycle
M phase: Mitosis; cell division, chromatids separate to daughter cells (1hr)
G1 phase: Gap; DNA of chromosomes present as single linear double helix of DNA; 10hrs
[G0: cells stopped dividing]
S phase: DNA Synthesis; 9hrs
G2 phase: Gap 2; Each chromosome has two identical sister chromatids; 4hrs
Describe the stages of Mitosis
Interphase- chromosomes not visible
Late Prophase- condensed chromosomes, each contains two sister chromatids
Metaphase- condensed chromosomes aligned in central plane of spindle
Anaphase- sister chromatids pulled to spindle poles
Telophase- Sister chromatids move to opposite poles of spindle
Cytokinesis- division of cytoplasm
