15 DNA and the Gene: Synthesis and Repair Flashcards
Describe the Hershey-Chase experiment
1) Label viruses: grow some viruses in the presence of P-32 for radioactive DNA and some with S-35 for radioactive protein.
2) Infect bacteria: Allow viruses with labelled DNA to infect a culture of E. coli cells.
3) Agitate Cultures: in kitchen blenders to separate virus capsules from bacterial cells.
4) Centrifuge Solution: to force bacterial cells into a pallet.
5) Record: location of radioactive labels.
What is DNA?
Deoxyribonucleic acid: a double helix molecule compose of four varying nucleotides with a deoxyribose sugar; that encodes for genetic information
What were the results and conclusion of Hershey-Chase experiment?
Results:
1) radioactive DNA is in pallet.
2) radioactive protein is in solution.
Conclusion: viral genes consist of DNA
Who proposed the double helix structure of DNA?
Watson and Crick
What does each deoxyribonucleotide consist of?
1) phosphate group
2) deoxyribose sugar
3) nitrogenous base
How do nucleotides link together?
Link together into a polymer by forming diester bond between the hydroxyl group on the 3’ carbon and the phosphate group on the 5’ carbon
What are the four nitrogenous bases?
Adenine
Thymine
Guanine
Cytosine
What are the base pairs?
Adenosine - Thymine
Guanine - Cytosine
What are the three alternative hypothesis about DNA synthesis?
1) Semiconservative replication
2) Conservative Replication
3) Dispersive Replication
What is the Semiconservative replication theory?
If the old, parental strands of DNA separated, each could then be used as a template for the synthesis of a new, daughter strand.
Each new daughter DNA molecule would consist of one old strand and one new strand.
What does the Conservative replication theory state?
If the bases temporarily turned outward so that complementary strands no longer faced each other, they could serve as a template for the synthesis of an entirely new double helix.
Would result in an intact parental molecule and a new daughter molecule consisting of entirely newly synthesized strands
What does the dispersive replication theory state?
If the parental double helix were cut wherever one strand crossed over another and DNA was synthesized in short sections by extending each of the cut parental strands to the next strand crossover, then there would be a mix of new and old segments along each replicated
Stretches of old DNA would be interspersed with new DNA down the length of each daughter cell.
What is the Meselson-Stahl Experiment?
An experiment that was used to determine how DNA is synthesized. Choosing between Semiconservative, conservative and dispersive replication.
Outline the process of the Meselson-Stahl Experiment
1) grow E. colin cells in medium with N-15 as a sole source of nitrogen for many generations. Collect sample and purify.
2) transfer cells to medium containing N-14. After cells divide once, collect sample and purify DNA.
3) after cells have divided a second time in N-14 medium, collect sample and purify.
4) centrifuge the three samples separately. Compare the locations of the DNA bands in each sample.
Outline the predictions of the Meselson-Stahl experiment
Predictions, after 2 generations:
- Semiconservative: 1/2 low density, 1/2 intermediate-density DNA
- Conservative: 1/4 high density DNA, 3/4 low density DNA
- Dispersive: all intermediate density DNA (hybrid)
Outline the results and conclusion of the Meselson-Stahl experiment
Results: after two generations 1/2 were low density (N-14) and 1/2 were intermediate density DNA (hybrid)
Conclusion: data from generation 1 conflict with conservative replication hypothesis. Data from generation 2 conflict with dispersive replication hypothesis. Therefore, replication is Semiconservative
In what direction can DNA polymerase add deoxyribonucleotides to DNA?
Can only add to the 3’ carbon, therefore, 5’ –> 3’ direction
Is DNA synthesis endergonic or exergonic?
Most polymerization reactions are endergonic (require energy input), however, DNA synthesis is exergonic (it releases energy).
The monomers that are used in DNA synthesis are deoxyribonucleoside triphosphates (dNTP). The release of the phosphate groups release enough energy for the reaction to occur.
DNA is ————- — that is, it occurs in both directions at the same time
Bidirectional. Therefore, replication bubbles grow in two directions as DNA replication proceeds.
Bacterial chromosomes have a —— —— of replication
Single origin
Eukaryotic chromosomes have ——– ——- of replication
Multiple origins
How is the helix opened and stabilized?
1) the enzyme DNA helicase breaks the hydrogen bonds between base pairs causing the two strands to separate
2) single-strand DNA-binding proteins (SSBPs) attach to the separated strands and prevent them from snapping back into a double helix.
3) unzipping process creates tension because of twisting forces
4) the enzyme, topoisomerase, cuts DNA, allowing it to unwind, and rejoins it ahead of the advancing replication fork. - releases tension.
What is a primer?
- A strand of a few nucleotides long that is bonded to the template strand
- provides DNA polymerase with a free 3’ hydroxyl group that can combine with an incoming deoxyribonucleotide to form phosphodiester bond.
How is the leading strand synthesis?
1) an enzyme primase synthesizes a short stretch of RNA that acts as a primer to DNA polymerase.
2) DNA polymerase begins working in a 5’ —> 3’ direction and adds deoxyribonucleotides to the complementary strand
3) as the polymerase moves along the DNA molecule, a doughnut-shaped molecule, called the sliding clamp, holds the enzyme in place on the template strand.
4) the enzymes product is called the leading strand, or continuous strand, because it leads into the replication fork and is synthesized continuously.
