Protoplasting and cell wall regeneration of the filamentous fungus aspergillus oryzae strain S1

Abstract

Aspergillus oryzae is well-known as an important filamentous fungus in food ferment and beverage industry in Japan. Its long standing application in beverage and industry has garnered its status as a GRAS (Generally Regarded as Safe) organism. availability of molecular genetic technology, where protoplasts serve as the pr! ingredient, offers the possibility of new routes to strain improvement and thus, ge manipulation of A. oryzae. In this study, a protoplasting method was conducted to a high yield of pro top lasts from A. 01J1zae, using an enzymatic approach. Several fac which contribute to high protoplast yield and protoplast cell wall regeneration studied. Results showed that the highest protoplast concentration obtained was 8.19 x protoplasts/ml, using 3 g wet-weight of a 20-hour mycelial culture harvested modifiedpeptone dextrose (PD) medium in a lytic mixture of 70 mg/ml Trichod. harzianum enzyme incubatedfor 4 hours in 0.8 M magnesium sulphate with dithio (DTT) pretreatment and 0.6 M sorbitol used as the solution for the separatio protoplasts. The highest frequency of protoplast regeneration obtained in this sIlt 65.70% using semi-solid Czapek regeneration medium with an agar concentration 0 %.