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KMID : 0380020100250020155
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Korean Journal of Biotechnology and Bioengineering
2010 Volume.25 No. 2 p.155 ~ p.166
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Strain Improvement through Protoplast Formation and Mutation of Inonotus obliquus Mycelia for Enhanced Production of Innerpolysaccharides (IPS) in Suspended Mycelial Cultures
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Hong Hyung-Pyo
Jeong Yong-Seob
Chun Gie-Taek
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Abstract
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Studies on the production of cell-wall bound innerpolysaccharides (IPS) (soluble β-D-glucan) have been performed by use of suspended myelial cultures of Inonotus obliquus. This product has promising potentials as an effective antidiabetic as well as an immunostimulating agents. As a first step to enhanced production of IPS, Intensive strain improvement programs were carried out by obtaining a large amounts of protoplasts for the isolation of single cell colonies. Rapid and large screening of high-yielding producers was possible because about fivefold higher amount of protoplasts (2.3 x 106 protoplasts/mL) could be recovered with relatively high regeneration rates of 10-2~10-3 by applying a modified filtration method, as compared to the previously used trapping method. A basic protocol necessary for UV-mutation of the protoplasts was also developed, resulting in several overproducing variants with good fermentation properties. Since the amount of IPS extracted from the mycelial cell walls of I. obliquus turned out to be almost constant per g DCW, increase in cell mass was considered the most important factor for the enhancement in IPS production. Therefore, attempts were made to screen mutant cells showing rapid mycelial growth rate in the final suspended cultures. Notably, the mutant strains showing an active cellgrowth in the preceding solid growth cultures were observed to produce higher amount of IPS in the suspended fermentations as well. A striking mutant, OBLQ756-15-5 strain, obtained from the survivors of a harsh UV-treated condition (97% death rate) was found to stably produce as high cell mass as 22 g DCW/L in the final fermentations. Currently, this strain is being tested for development of a scaled-up fermentation process for mass production of IPS.
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KEYWORD
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Inonotus obliquus, strain improvement, protoplasts, mycelia, innerpolysacchrides (IPS), β-D-glucan
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