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The evacuolated protoplasts were suspended in three volumes of TR buffer (30 mM HEPES-KOH (pH 7.4), 80 mM potassium acetate, 0.5 mM magnesium acetate, 2 mM DTT) supplemented with one tablet per 50 ml of Complete EDTA-free Protease Inhibitor Mixture (Roche Diagnostics, Mannheim, Germany) and disrupted using the nitrogen decompression method in a cell disruption vessel (Parr Instrument, Frankfurt, Germany) for 30 min at 10 bar

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When the grinding is done, pop on the screw-on top and store your ground herb away