BIO 344 1st Edition Lecture 4Outline of Last Lecture I.DNA Polymerizationa. Chemistry of deoxy and ribonucleoside triphosphatesII.Fidelityb. ProofreadingIII. ChemotherapeuticsIV. Dideoxyribonucleoside triphosphateV. “Seeing DNA”—Technologya. Radioactive labelingb. Fluorescent labelingVI. DNA Sequencinga. Sanger Methodb. Gel Electrophoresisc. PCRd. NextGenOutline of Current Lecture I. Analyzing papers-- example: Meselson Stahl Papera. Question: mechanism of replicationb. Hypothesisc. Data and resultsII. The Meselson Stahl experimenta. N15 and N14b. conclusionCurrent LectureHow to analyze papers—example: Meselson Stahl Paper- determine the question:These notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.o what is the mechanism of replication? Conservative, semiconservative, or random dispersive- Determine their hypothesiso Isotopic labels can be used to determine the method of replication- Data and resultso Presented data should support hypothesiso Look at and interpret data in context with the experiment- Resultso Read and interpret the figureso Think about what their expected result was and if that expectation is correctThe Meselson Stahl Experiment- N15: more neutrons, heavier than N14 (the natural nitrogen found in DNA nucleotides)o Grew cells in N15 for many generations so that all proteins, bases were labeled with N15 (heavier than natural) then transferred to medium with N14 Fed bacteria ammonium chloride to incorporate N14—but how does is become part of their DNA?- Add precursors for DNA synthesis (ribosides of nucleotides with N14)- Graph shows rate of replication and change between nitrogen shif- Shows that cells are not affected by change between N15 and N14o Doing this to eliminate confounding factors Collect cells to determine density of DNA—did they actually use N14?- Density centrifugation to separate molecules by weighto See slide for illustration- N14 bond will be higher sedimented than N15- We can see where DNA is by shining UV light and areas of absorption create bandso As generations progress, you see a shif in bands (to the lef—lower density), and eventually two bands shifed to the lef and eventually just one band to the far lef Interpret as: “new DNA” is labeled with N14 and original is N15—semiconservative is likely the method- Meselson ans Stahls Conclusion:o Not random dispersion replication—more big questions need to be
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