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Today’s Outline. Study Notes 6b Due! Take up exam View exam on Friday between 2-4pm in C-4. Written appeals-process DNA Replication Replication PCR Telomeres DNA Replication Tutorial 3-5pm Friday. Write Down:. What is one thing you learned about DNA replication from the readings?. - PowerPoint PPT PresentationTRANSCRIPT
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Today’s Outline
• Study Notes 6b Due!• Take up exam – View exam on Friday between 2-4pm in C-4.– Written appeals-process
• DNA Replication– Replication– PCR– Telomeres
• DNA Replication Tutorial 3-5pm Friday
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Write Down:
What is one thing you learned about DNA replication from the readings?
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To Understand DNA Replication, Understand the Parts
- Helicase- Topoisomerase- Single Strand Binding Proteins- DNA Polymerase III- Sliding Clamp- DNA Primase
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HelicaseHelicase unzips DNA.
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TopoisomeraseRelieves the tension produced when DNA is
unzipped.
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Single Strand Binding Proteins
Stabilizes DNA and prevents it from binding to parallel strand.
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DNA Polymerase III
Adds dNTPs to the newly synthesized strand on the leading strand. Adds in a 5’ 3’ direction.
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Sliding Clamp
Holds DNA Polymerase III in place.
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DNA Primase
Nucleotides can ONLY be added to the 3’ end of an already-attached nucleotide. A primer
provides this. DNA Primase builds the primer.
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DNA
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Polymerase Chain Reaction
• What does polymerase do?• What if….– I put an entire genome in a test tube.– I added DNA polymerase III and dNTPs.– I added very specific DNA primers that mapped
onto a specific gene.– Then I started heating and cooling the DNA over
and over and over and over….
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Polymerase Chain Reaction5’
5’
3’
3’
5’
3’
5’3’
5’
3’
5’ 3’
- taq polymerase- dNTPs- primers- buffer
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Discuss:
• Find three similarities and three differences between (i) DNA replication on the leading strand and (ii) DNA replication via PCR.
(Debrief using name cards)
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The 1st Challenge of Lagging Strand Replication
• DNA Primase is used over and over• DNA Primase lays down ribonucleic acid.
Solution: an enzyme that can replace RNA nucleotides with DNA nucleotides DNA Pol1
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The 2nd Challenge of Lagging Strand Replication
Solution: an enzyme that can connect 3’ carbons to phosphate groups
LIGASE
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Ends of parental DNA strands
Leading strandLagging strand
Lagging strand
Last fragment Previous fragment
Parental strand
RNA primer
Removal of primers and replacement with DNA where a 3 end is available
Second round of replication
New leading strand
New lagging strand
Further rounds of replication
Shorter and shorter daughter molecules
5
3
3
3
3
3
5
5
5
5
53
DNA Pol1 Can only add nucleotides to the 3’ end of existing DNA.
Telomeres protect an organism’s genes from being eroded during successive rounds of DNA replication. In humans the sequence TTAGGG is repeated between 100 and 1000 times.
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Which of these replaces RNA with DNA?
A) DNA Polymerase 1B) DNA Polymerase 3C) DNA PrimaseD) LigaseE) Nuclease
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Which of these glues 3’ and 5’ DNA ends together?
A) LigaseB) DNA Polymerase 1C) Binding ProteinsD) DNA Polymerase 3E) DNA Primase
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Which of these adds new nucleotides to the leading strand?
A) DNA Polymerase 3B) The Sliding ClampC) DNA Polymerase 1D) DNA PrimaseE) Binding Proteins
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Which of these is not needed by the leading strand:
A) DNA Polymerase 1B) Binding ProteinsC) DNA PrimaseD) The Sliding ClampE) DNA Polymerase 3
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Which of these is not needed by the lagging strand:
A) The Sliding ClampB) DNA Polymerase 1C) Binding ProteinsD) DNA Polymerase 3E) All are needed
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Why are telomeres needed?
a) Primers can’t bind to 5’ ends of DNAb) DNA polymerase 1 can’t add nucleotides to 5’
ends of DNAc) Okazaki fragments don’t form at the end of
DNA strandsd) Helicase can’t bind to the end of DNA
strands.e) None of the above.
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Write Down:
1. On a scale of 1 to 10, how well do you understand DNA replication?
2. What is one thing you understand really well?
3. What is one thing you do not understand?
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Coming Up:1. Sanger Sequencing (Tues)2. Molecular Sculpting of DNA replication (Tues)3. Bonus assignment due (see Lecture Schedule)4. Field trip sign up5. DNA Replication Tutorial 3-5pm Friday