Rapid Isolation of Nuclei from Carrot Suspension Culture Cells Using a BioNebulizer

The BioNebulizer is an instrument that breaks cells and large molecules using the shearing forces created by laminar flow of high-pressure gas in the microcapillary channels generated by the instrument. Within 4 min, 90% of the carrot suspension culture cells that passed through the nebulizer were broken. Cytosol and organelles were released from the broken cells leaving cell wall ghosts. Nuclei were further purified by means of a discontinuous Percoll gradient. This method yielded an average of 2 x 10(5) nuclei from 2 g of suspension culture cells (approximately 2 x 10(6) cells). The isolated nuclei actively incorporated [8,5'-3H]-GTP into RNA.