1990 Fiscal Year Final Research Report Summary
Establishment of Protoplast Regeneration Method for Efficiently Dedikaryotizing Higher Basidiomycetes
| Project/Area Number |
63860006
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| Research Category |
Grant-in-Aid for Developmental Scientific Research (B).
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| Allocation Type | Single-year Grants |
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| Research Field |
Breeding science
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| Research Institution | The Japan Kinoko Research Center Foundation |
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Principal Investigator |
KOMATSU Mitsuo The Japan Kinoko Research Center Foundation, The Tottori Mycological Institute, Chief of Section, 菌蕈研究所, 研究部長 (60088846)
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| Co-Investigator(Kenkyū-buntansha) |
MAETA Yoshiyuki The Japan Kinoko Research Center Foundation, The Tottori Mycological Institute,, 研究員 (90209370)
SATO Fumihiko Kyoto University, Faculty of Agriculture, Assistant professor, 農学部, 助手 (10127087)
NAKAI Yukitaka The Japan Kinoko Research Center Foundation, The Tottori Mycological Institute,, 研究室長 (00088840)
YAMADA Yasuyuki Kyoto University, Faculty of Agriculture, Professor, 農学部, 教授 (50026415)
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| Project Period (FY) |
1988 – 1990
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| Keywords | Higher basidiomycetes / Protoplast regeneration / Contribution to mushroom strain improvement / 03Establishment of an efficient dedikaryotization method |
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| Research Abstract |
In the higher basidiomycetes, dissociation of dikaryons into the component monokaryons (= neohaplonts), namely dedikaryotization, is expected to offer a significant opportunity for studying nucleo-cytoplasmic relation and somatic recombination as well as strain improvement and has been repeatedly carried out by artificial means such as microsurgical operation and chemical treatment. However, these methods are not desirable because of the reason that they need skillful and time consuming work, and often produce neohaplonts with only one type of two component nuclei. In this study, we therefore investigated the possibility of dedikaryotization via protoplast formation and regeneration in ten edible mushroom species, including Lentinus edodes and Pleurotus ostreatus, to establish a new method suitable for dedikaryotizing a variety of higher basidiomycetes with ease. The results are summarized as follows.
Conditions for efficient production and regeneration of mycelial protoplasts from dikaryons of all mushroom species used were developed. When the isolated protoplasts were incubated at 25C in regeneration agar medium, about 3 to 40% of them started regeneration into hyphae within 3 days and formed visible colonies, varying in size, after 7 - 10 days of incubation. By isolating preferentially small colonies out of these colonies reverted from protoplasts, it was found that neohaplonts could be recovered at the high frequency of 30 - 95% in all mushroom species used and also the two component nuclear types appeared.
Although the neohaplonts obtained showed a wide range of mycelial growth rates, the good-growing ones exhibited no variation in biological properties such as colony morphology and electophoretic zymograms of esterase and malate dehydrogenase from the parental monokaryons. From these results, it is concluded that the protoplast regeneration method for dedikaryotizing higher basidiomycetes developed in this study is more reliable and useful than previous methods.
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