Figure 1

Outline of the procedures used for isolation of nuclei. A) Species-specific adjustments at specific steps of isolation of nuclei from the leaves of tobacco, potato and apple. NIB, buffer for isolation of nuclei. B) Isolation of nuclei using Percoll/sucrose density gradient centrifugation (step 4) shown in more detail. The crude preparation of nuclei is loaded on the top of 60% Percoll/2.5 M sucrose density gradient before centrifugation. After centrifugation (1000 × g for tobacco and potato, 1200 × g for apple), the Percoll layer contains most of the nuclei (see Nuclear fraction) and is collected for further purification on 35% Percoll cushion. Also the two other fractions (60% P/2.5 M S interface and 2.5 M Sucrose layer) were subjected to western blot analysis (see Figure 2). The shaded fraction on the bottom of the tube after centrifugation contains mainly starch grains.