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Sedimentation etc

Explore the general principles of sedimentation in biophysics, including density, sedimentation coefficient, and boundary determination. Learn about analytical centrifuges, diffusion, and density gradient separation techniques. Discover electrophoresis methods for DNA and hemoglobin separation.

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Sedimentation etc

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  1. Sedimentation etc Topic 3 Part 1 Biophysics

  2. General Principles f v Sphere: f0 = 6phR F (like mg) Other particle: get r = f/ f0 and f = r f0 Example – prolate with a = 3 and b = 2

  3. Sedimentation 1- r is the buoyancy factor ~ 1- ro/r (if ro > r then it floats). s = sedimentation coefficient, [s] = Svedberg, 1 x 10-13 sec = 1 Svedberg. Density of medium, r = mo/V, V = volume. is specific volume, volume/mass of substance in solution (V/m)

  4. Determining sAnalytical Centrifuge This instrument scans absorption along the centrifuge cell as a function of time – giving concentration.

  5. Determining s vb = drb/dt = w2srb, where the subscript b signifies the boundary (so rb is the boundary between solvent and solution). • So plot ln(rb) vs t and get slope which is equal to w2s.

  6. Determining sDiffusion blurs boundary D = RT/NAf,

  7. More on s • Depends on temperature and viscosity so define for standard conditions • s depends on M and f (shape) so if know M can get f (shape information/stokes radius)

  8. Density Gradient used for separation

  9. Real data on hemoglobin free and bound to haptoglobin

  10. Electrophoresis Now have F = ZeE, fv = ZeE. Mobility, U = v/E = Ze/f Sphere: U = Ze/(6phR)

  11. Movement on a Gel • This is a gaussian centered around x = xo + mFt • With rms of , velocity = mF, • m = 1/(6phr) with r = radius and h = viscosity • D = mkBT = diffusion constant

  12. Running DNA on a Gel • Closed small plasmids give discrete bands • Long DNA tunnels and separation goes as 1/mass • Can get screening from lots of cations • 2-D electrophoresis good for large pieces of DNA • Genome project used Saenger method

  13. Running Plectonemic helices • L = T + W • For DNA at several Kbp, helices run by writhe and hence (for constant twist), L • Topoisomers for DNA of defined length give gaussian band due to different energies • Wang Paper http://www.cbs.dtu.dk/staff/dave/roanoke/supercoil.jpg

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