Y. Fukumasanakai et al., DEDIKARYOTIZATION OF THE SHIITAKE MUSHROOM, LENTINULA EDODES BY THE PROTOPLAST REGENERATION METHOD, Journal of General and Applied Microbiology, 40(6), 1994, pp. 551-562
The present study was carried out to investigate the applicability and
usefulness of the protoplast isolation and regeneration method as a n
ew technique for artificially dedikaryotizing Lentinula edodes dikaryo
ns. When protoplasts derived from dikaryans were incubated in a regene
ration agar medium at 25 degrees C, about 11% of them individually sta
rted regeneration into hyphae within 3 days and formed visible colonie
s, varying in size, after 7 days of incubation. By isolating preferent
ially smaller colonies out of these visible colonies, it was found tha
t neohaplonts could be obtained at the high frequencies of 40-92% in e
very dikaryon sampled and that the two-component nuclear types appeare
d. These neohaplonts showed considerable variation of mycelial growth
rates. However, robust neohaplonts exhibited no apparent change in bio
logical properties such as colony morphology, and electophoretic zymog
rams of esterase and malate dehydrogenase, suggesting that they might
retain the original genetic traits. From these results, it is conclude
d that the protoplast regeneration method for dedikaryotizing L. edode
s dikaryons is more useful than previous methods such as the physical
procedure and chemical treatment.