Table 2 Summary of protoplast preparation parameters for some common fungal species
From: Methods for genetic transformation of filamentous fungi
Strain | Starting material | Enzyme | Enzymolysis conditions | Osmotic stabilizer | Protoplast yield (protoplasts/mL) | References |
|---|---|---|---|---|---|---|
A. niger K10 | NA | Lyophilized snail | Incubate at 31 °C for 3 h on a laboratory reciprocal shaker (78 strokes/min, amplitude 3 cm) | 0.7 M NaCl | 8.9 × 106 | [31] |
A. niger N583 | Mycelium | Lysing enzymes, chitinase griseus and β-glucuronidase | Incubate in a 100 mL glass bottle for 2 h at 37 °C and 130 rpm | 0.8 M sorbitol | NA | [32] |
Aspergillus nidulans A89 | Mycelium | Yatalase, Kitalase | Incubate 6 h at 30 °C with constant shaking (60–80 rpm) | 1 M sorbitol | NA | [33] |
Aspergillus nidulans G191 | Mycelium | Novozym 234 | Incubate 1.5 h at 30 °C | 0.6 M KCl | NA | [34] |
Trichoderma atroviride T11 | Mycelium | Lysing enzymes from Trichoderma. harzianum | Incubate at 30 °C for 2 h with shaking at 80 rpm | 1 M sorbitol | 1–5 × 106 | [26] |
Penicillium simplicissimum | Mycelium | Driselase, caylase, cellulase, enzyme cocktail II from Merck company | Incubate at 30 °C for 20 or 180 min | 1.2 M sorbitol | 2 × 109 | [26] |
Trichoderma reesei QM 9414 | Mycelium | Lysing enzymes from T. harzianum | Incubate at 28–30 °C for 2 h | 1 M sorbitol | NA | [17] |
Trichoderma reesei QM 9414 and VTT-D-79125 | Mycelium | Novozym 234 | Incubate at 28 °C for approx. 1.5 h | 1 M and 1.2 M sorbitol | 5 × 107 to 5 × 108 | [16] |
N. crassa M246 (qa-2 mutation) and M6-11 (arom-9 mutation) | Mycelium | Proteinase K | Incubate at 1 h at room temperature | 1 M sorbitol | 1 × 107 | [19] |
Aspergillus fumigatus 2085 | Mycelium | β-glucuronidase | Incubate at 37 °C for 1 h with occasional shaking | 0.7 M KCl | 1 × 106 | [35] |
A. oryzae NRRL 492 | Mycelium | Novozyme 234 and β-glucuronidase | Incubate at 30 °C for 1.5 h with shaking at 80 rpm | 0.6 M sorbitol and 1 M sucrose | Each 400 mL culture yielded 5–10 × 107 protoplasts | |
Rhizopus oryzae AS 3.819 | Mycelium | Helicase, cellulase, lyticase | Incubate at 35 °C for 140 min with gentle shaking | 0.6 M sorbitol | 1 × 107 | [36] |
Monascus purpureus | Mycelium | Glucanex, lysing enzyme from T. harzianum β-glucuronidase and Caylase | Incubate at 28 °C for 4 h with orbital shaking (100 rpm) | 0.5 M KCl and 0.1 M MgSO4 | 1 × 109 | [22] |
Penicillium expansum | Mycelium | 0.6% cellulase and 0.6% snailase | Incubate at 25–35 °C for 3–3.5 h | 0.6 M NaCl | 2.36 × 108 | [37] |