Isolation of Nuclei from a Tetraploid Strain of Saccharomyces cerevisiae

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Abstract
A procedure was developed for isolating nuclei from cells of a tetraploid strain of Saccharomyces cerevisiae. In this procedure, spheroplasts, prepared from young yeast cells using the two bacterial glucanases, were lysed (a) at low osmotic pressure in the presence of a detergent or (b) at a moderate osmotic pressure in the absence of detergent, and the lyzate was homogenized and fractionated by centrifugation through discontinuous sucrose gradients. This procedure gave purified nuclei in 20—30% yield. The nuclei isolated using condition (a) for lysis were devoid of a nuclear envelope, while those isolated using condition (b) retained their envelopes. The nuclei isolated using either of the conditions of lysis had two to three times more RNA than DNA, and retained RNA polymerase [EC 2.7. 7.6] and NAD pyrophosphorylase [EC 2.7.7.1] activities.

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