Abstract
A novel method for isolatingEuglena gracilis Z. nuclei, based on pretreatment of cells in concentrated glycerol buffer before homogenization, is described. Such a treatment weakens the tough cell pellicle facilitating cell disruption, and avoids nuclear damage induced by detergents and by freezing and thawing the cells in aqueous media. Nuclei, purified by centrifugation in dense sucrose, are obtained with a 30% yield, and only small amounts of cell wall fragments contaminate the nuclear pellets. The purified nuclei retain their ultrastructural characteristics. High molecular weight DNA, as well as undegraded RNA species and histones, can be extracted from these nuclei. Nuclease digestions and spread preparations show an unaltered nucleosomal structure of chromatin. This method has been applied to cell samples at any stage of the cell cycle, including mitosis, since inEuglena the nuclear envelope persists during cell division.
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