Sedimentation velocity of DNA in isokinetic sucrose gradients: calibration against molecular weight using fragments of defined length.

The relationship between sedimentation coefficient and molecular weight for DNA sedimenting in preformed alkaline and neutral sucrose gradients was determined using absolute molecular weight standards (restriction fragments of plasmid pBR322 and phage lambda DNA). The range of calibration for alkaline gradients was extended to small DNA fragments (652 base-pairs) for the first time. The exponent b in the equation S20 degrees, w = aMb was found to be 0.380 in neutral gradients and 0.410 in alkali. The latter value differs significantly from previous estimates. The gradients were isokinetic, and the distance sedimented was shown to be directly proportional to the sedimentation coefficient at all times.