Abstract
The usual method of alkaline (0.1-0.5M NaOH) sucrose density gradient sedimentation of intact cells enables DNA molecular weight determinations with a minimum of shearing from bacterial protoplasts. This technique however has not yet been in general successfully applied to intact mammalian cells because of higher molecular weight and contamination by protein, causing the DNA to sediment in a nonregular fashion. In addition, DNA aggregates may form which sediment to the bottom of the tube causing low recovery of the layered radioactive counts. This problem has been overcome for Ehrlich ascites cells, by including 6M guanidinium chloride in the alkaline sucrose density gradient made 0.9M in sodium hydroxide. By this method, virtually complete recovery of the radioactivity layered upon the gradient is obtained in the recovered fractions, along with a reproducible DNA peak position. Neutral sucrose gradients of Ehrlich ascites cell DNA are also possible if 6M guanidinium chloride is included in the gradient. ...
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