Question

JO 8-9 In the classic paper that demonstrated the semi-con- servative replication of DNA, Meselson and Stahl began by showing that DNA itself will form a band when subjected to equilibrium sedimentation. They mixed randomly frag- mented E. coli DNA with a solution of CsCl so that the final solution had a density of 1.71 g/ml. As shown in Figure Q8-2, with increasing length of centrifugation at 70,000 times gravity, the DNA, which was initially dispersed throughout the centrifuge tube, became concentrated over time into a discrete band in the middle A. Describe what is happening with time and explain why the DNA forms a discrete band. Egge hours centrifugal field - the the tere car end Figure 28-2 Ultraviolet absorption photographs showing successive stages in the banding of E.coli DNA (Problem 8-9). DNA which absorbs UV light, shows up as dark regions in the photographs. The bottom of the centrifuge tube is on the right. (From M. Meselson and F.W. Stahl, Proc Natl Acad Sci USA 44:671-682, 1958. With permission from National Academy of Sciences frag 8-1 mus thes B idea mo hun
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Answer 1

This is the classic Meselson and Stahl experiment which demonstrated the semi-conservative mode of replication in DNA.

A. With time, the rate of sedimentation of DNA molecules approaches the rate of diffusion of DNA in the given solution. When both these opposing forces balance at equilibrium, the DNA molecules with a particular molecular weight (MW) assemble together in a band that narrows with time. At this point, the buoyant density of the DNA molecules becomes equal to the density of the solution in which they are suspended.

B. As such, the buoyant density of the DNA in this case will be equal to the density of CsCl solution, which is 1.71 g/mL.

C. There is a limit to the concentrating tendency of DNA molecules of similar MW, owing to opposing diffusion forces that distribute it over a band in an inverse relation to its MW.