BIOCHEMISTRY QUESTION 5. In 1958, Meselson and Stahl published an elegant experi
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Question
BIOCHEMISTRY QUESTION
5. In 1958, Meselson and Stahl published an elegant experiment showing how double-stranded DNA replicates in E. coli.
-DNA was uniformly labeled with a heavy (H) atom of nitrogen by growing cells for 14 generations with N15H4Cl as the sole nitrogen source.
-N15 cells were diluted into medium containing light (L) N14 medium.
- Cells were grown for multiple generations.
- At specific times, DNA was isolated and centrifuged to equilibrium in a CsCl gradient diagrammed on the right. L/L, L/H, and H/H DNA have distinct boyant densities in CsCl gradients.
- DNA was detected by A260 from gradient fractions.
If DNA replicates semi-conservatively (see Figure 33.15 and 16),
A. Would there be any L/L DNA after one generation of growth?
B. How much H/L would there be after one generation?
C. If DNA replicated conservatively, what would be detected after 1 generation?
6. Below are two different DNA replication patterns that could have been detected by Meselson and Stahl.
Label each type of replication pattern below the diagram.
Alongside the diagram, for each type of replication pattern, give the expected percentages of Heavy (black), Light (gold), and HL (black and gold duplex) DNA molecules after 1 (F1) and 2 (F2) replication cycles in light medium. Recall that cells initially were grown in heavy medium (15NH4Cl) for many generations before shifting to light medium (14NH4Cl).
Explanation / Answer
5).
(A) No
(B) All the DNA after first generation will be in H / L form.
(C) If DNA replicates conservatively L / L and H / H will be in equal proportion.
6).
Figure on left (Conservative DNA replication)
F1: 50% H / H duplex and 50% L / L duplex
F2: 25% H / H duplex and 75% L / L duplex
Figure on right (semi-conservative DNA replication)
F1: 100% H / L duplex
F2: 50% H / L duplex and 50% H / L duplex.
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