the enzyme/protoplast solution was
filtered through a 35-lm nylon mesh. The filtered solution was
centrifuged in a round-bottomed tube for 5 min at 100g to pellet
the protoplasts. The protoplasts were resuspended in 10 mL cool
W5 solution (2 mM MES, pH 5.8, 154 mM NaCl, 125 mM CaCl2,
5 mM KCl, and 5 mM glucose) and then centrifuged at 100g for