Protoplast Isolation and Culture

Protoplasts are cells of plants, fungi, or bacteria in which the cell wall has been removed, but the plasma membrane is intact. Among the many uses for these “naked” cells is transient or permanent genetic transformation by introduction of trans-gene DNA, somatic hybridization by protoplast fusion of species or subspecies resistant to traditional cross-breeding, and isolation of sub-cellular organelles. We offer a number of enzyme products to break down the cell wall. Individual hydrolytic enzymes include cellulases, hemicellulase, pectinase, and pectolase. Some, such as pectinase from Rhyzopus sp. or DriselaseTM are crude extracts with several specificities, while two of our thermostable cellulases are nearly pure recombinant preparations. Listed also are several reagents necessary for handling protoplasts, including the vitamin mixture for Kao and Michayluk culture medium, fluorescein diacetate for determinate of protoplast viability, and a specially formulated plant protoplast digest/wash solution.

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C1184 Cellulase from Aspergillus niger powder, ≥0.3 units/mg solid
C1794 Cellulase from Trichoderma sp. BioReagent, plant cell culture tested, 3-10 units/mg solid
C9499 Cellulase, thermostable from Clostridium thermocellum recombinant, expressed in E. coli, ≥90% (SDS-PAGE)
C9624 Cellulase, thermostable from Dictyoglomus turgidum recombinant, expressed in E. coli, ≥90% (SDS-PAGE)
D8037 Driselase Basidiomycetes sp. plant cell culture tested, BioReagent
F7378 Fluorescein diacetate used as cell viability stain
F7250 Fluorescein isothiocyanate isomer I suitable for protein labeling, ≥90% (HPLC), powder
F3543 Fluorescent Brightener 28 used as a stain and brightening agent
G5768 Gamborg’s B-5 Basal Salt Mixture fine powder, plant cell culture tested
H2125 Hemicellulase from Aspergillus niger powder, 0.3-3.0 unit/mg solid (using a β-galactose dehydrogenase system and locust bean gum as substrate)
K3129 Kao and Michayluk Vitamin Solution 100 ×, liquid, suitable for plant cell culture
P0690 Pectinase from Aspergillus niger BioReagent, plant cell culture tested, aqueous glycerol solution, ≥5 units/mg protein (Lowry)
P2401 Pectinase from Rhizopus sp. powder, 400-800 units/g solid
P5936 Pectolyase from Aspergillus japonicus plant cell culture tested, lyophilized powder
P7828 Percoll® plant cell culture tested
D9692 Plant Protoplast Digest/Wash Solution Rapid isolation of viable protoplasts from plant tissue
P2307 Polyvinylpyrrolidone plant cell culture tested, average mol wt 10,000