• 한국균학회소식:학술대회논문집
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• 한국균학회 2009년도 추계학술대회 및 정기총회
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• pp.14-16
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• 2009

Poria cocos is an importantant medical macrofungus,the sclerotium of Poria cocos has specific value as the drug material. There are few papers about its breeding and spawn rejuvenation. In this project, the protoplasts of cultivated strain T and wild strain L were prepared and treated separately by ultraviolet and heating, then fused with the PEG6000. The tural fusants were selected and identified by the affinity and ISSR analysis. 71 incompatibility strains between parents and reg regenerations were obtained from 118 regenerations by the affinity analysis. Five incompatibility strains were amplified with different primers, the results were showed that they had specific bands of both parents in the profile amplified with 3 primers, which proved these 5 strains were fusants by means of molecular biology marker. On the other hand, 25 strain were selected from 168 protoplast regenerations of cultivated strain T for cultivation experiment. The fresh sclerotium weight of these protoplast regenerations were better than the original strain.significantil 3 strains (T-1, T-4, T-7) increased respectively 118%, 73% and 73% than original strain. This method could be the effective in the rejuvenation Poria cocos.

• 한국미생물·생명공학회지
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• 제14권5호
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• pp.359-363
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• 1986

본 실험의 목적은 starch의 alcohol발효에 있어서 전분의 $\alpha$-1.4 linkage을 분해하여 alcohol발효를 하는 S diastaticus와 전분 중의 $\alpha$-1.4, $\alpha$-1.6 linkage를 모두 분해할 수 있는 C. tropicalis 간의 이속간의 protoplast fusion을 시켰다. 이때 세포 융합의 빈도는 $10^{-5}$-$10^{-6}$이였으며, 이틀 융합체중 amylase 생성능과 유전적으로 안정한 융합체를 분리하였다. fusant의 성질을 조사하기 위하여 탄소원 자화능을 조사한바 parent와 달리 탄소원 자화능이 일부 보완됨을 보였고 또한 fusant는 원 parent보다 세포의 크기가 클 뿐만아니라, DNA함량도 많았다. spore형성능은 S. diastaticus A4는 spore를 형성하나 C. tropicalis는 형성할 수 없는 반면에 fusant는 형성하였고, Cu$^{++}$ 내성과 NaCl 내염성도 조사하였는데 fusant는 parent의 중간 성질을 가졌다. Fusant의 증식율을 조사하기 위하여 생육도를 조사한 결과 Parent보다 유도기가 길었음을 알았다.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 1986년도 추계학술대회
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• pp.528.1-528
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• 1986

Cellulose utilising hybrids between Cellulomonas fimi and Bacillus pumilus were isolated after PEG mediated protoplast fusion. 33% (w/v) PEG #6, 000 and 50mM $Ca^{++}$were optimum concentration. The intergeneric fusion frequency was 3.2$\times$10$^{-7}$ xtracellular CMCase and $\beta$-glucosidase activities were detected from one hydrid unlike only CMCase was detected from Cellulomonas fimi.

• Mycobiology
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• 제34권2호
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• pp.73-78
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• 2006

Inter-specific hybridization between Pleurotus pulmonarius and P. florida was attempted through PEG-induced protoplast fusion to select a fusant. The protocol for protoplast release, regeneration and fusion in these two Pleurotus species was standardized using the variables controlling the process. The mixture of mycolytic enzymes, i.e. commercial cellulase, crude chitinase and pectinase, KCl (0.6 M) as osmotic stabilizer, pH 6 of the phosphate buffer and an incubation time of 3 hours resulted in the maximum release of protoplasts from 3-day-old mycelia of P. florida ($5.3{\sim}5.75{\times}10^{7}$ protoplasts/g) and P. pulmonarius ($5.6{\sim}6{\times}10^{7}$ protoplasts/g). The isolated protoplasts of P. florida regenerated mycelium with 3.3% regeneration efficiency while P. pulmonarius showed 4.1% efficiency of regeneration. Polyethyleneglycol (PEG)-induced fusion of protoplasts of these two species resulted in 0.28% fusion frequency. The fusant produced fruiting bodies on paddy straw but required a lower temperature of crop running ($24{\pm}2^{\circ}C$) than its parents which could fruit at $28{\pm}2^{\circ}C$. The stable fusant strain was selected by testing for the selected biochemical markers i.e. Carbendazim tolerance and utilization of the lignin degradation product, vanillin.

• 한국식품과학회지
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• 제25권4호
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• pp.366-372
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• 1993

본 연구는 미림 제조시 미림의 품질을 높이기 위한 방법의 하나로 세포 원형질체의 융합체 의해 ACPase 활성이 높은 균주를 얻으려고 시도하였다. 돌연변이 방법으로 acid carboxypeptidase(ACPase) 활성이 높은 A. oryzae 9-12와 A. shirousamii IFO 6082-60을 선별하여 사용하였다. A. oryzae 9-12와 A. shirousamii IFO 608260의 원형질체의 형성조건은 chitinase (10mg/ml), cellulase(10mg/ml) 및 zymolase(20T(5mg/ml)등의 효소 혼합용액에서 $30^{\circ}C$에서 4시간 반응시켰을 때 가장 높았으며, pH6.0일 때, 안정제로서는 0.6M KCl 및 무기 염류로서는 0.01M $CaCl_2\;2H_2O$가 가장 높았고, 30% PEG 6,000이 가장 좋았다. 이때의 융합빈도는 0.71%였다. 융합주의 ACPase의 활성은 F-50이 20,800unit/g으로 변이주보다 1.4배 높았다.

• Journal of Microbiology and Biotechnology
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• 제2권1호
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• pp.46-49
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• 1992

The availability of Phaffia rhodozyma as an astaxanthin sources in the aquaculture industry is limited because of the low carotenoid content of natural isolate. In this study, we have used the protoplast fusion technique to construct cell hybrids with an increased content of astaxanthin from P. rhodozyma. Cell hybrids (F307 and F406) obtained were very stable and produced considerably more astaxanthin (> 1 mg/g yeast) than the wild parent. Karyogamy was confirmed by the isolation of recombinants after mitotic segregation of parental auxotrophic genetic markers, the increased amount of chromosomal DNA/cell and the presence of single nucleus/cell.

• 한국자원식물학회지
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• 제19권3호
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• pp.418-421
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• 2006

In this study, we surveyed the influence of IOA (iodoacetamide) and media upon protoplast fusion for the efficient production of the somatic hybrid among S. sisymbriifolium and other Solanum species (S. intergrifolium and S. toxicarium). Regardless of a breed, as the IOA concentration increases, the cell division tends to decrease. As the influence of the media on the colony formation, we could get 5 colonies from the fusion of S. sisymbriifolium and IOA-treated S. integrifolium protoplast, but none was observed in other fusions. As a result of analyzing their IDH isozyme, we found a somatic hybrid in 2 objects.

• 미생물학회지
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• 제27권1호
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• pp.27-34
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• 1989

In order to evaluate the applicability of enzyme electrophoresis for the identification of intra/interspecific hybride obtained by the protoplast fusion in Trichoderma, soluble proteins, intracellular soluble enzymes and extracellular $\beta$-glucosidase were analyzed by polyacrylamide gel electrophorsis. As the results, patterns of soluble protein, and isozyme patterns of peroxidase, malate dehydrogenase, and $\beta$-glucosidase in hydrids were defferent from those in parental and wild type strains. Therefore, it was established that the analysis of protein pattern by electrophoresis could be applied to isolate and identify the hybrids from the protoplast fusion.

• KSBB Journal
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• 제5권3호
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• pp.255-261
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• 1990

Astaxanthin을 생산하는 효모 Phaffia rhodozyma로부터 제조된 상보적 돌연변이 균주간의 세포융합을 통하여 astaxanthin을 대량 생산하는 균주를 얻고자 시도하였다. 이들의 원형질체 융합빈도는 $1.3$\times$10^-^5-6.0$\times$10^-^5$이었고 DNA함량, 핵염색, UV조사에 대한 생존력 비교 그리고 형질분리분석등으로 핵융합을 확인하였다. 융합체 중 F1은 야생형의 모 균주와 비교했을때 astaxanthin생성량이 약 3배 증가하였다.

• 미생물학회지
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• 제28권4호
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• pp.311-317
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• 1990

Intraspecific and interspecific protoplast fusions in/between C. thermocellum and C. thermohydrosulfuricum were studied. Protoplast fusions were well induced in 30-40% PEG solution, however, their fusion frequencies were low level of 1.2*10$^{-7}$ for intraspecific fusion of C. thermocellum, $6.7*10^{-7}$ for C. thermohydrosulfuricum, and 4.2*10$^{-7}$ for interspecific fusion between above two Clostridia, respectively. Most fusants were unstable and segregated after 3 subcultures. Relatively stable intraspecific C. thermocellum fusant FTT17, intraspecific C. thermohydrosulfurecum fusant FSS22 and interspecific fusant FTS3, which were stable after several subcultures, were selected and properties of fusants were further investigated, Phenotypes of the fusants were similar with wild types mostly in cellular morphology, carbon source assimilation and enzyme activities. However they were differed in assimilation of pyruvic acid and sorbitol as carbon source. The DNA contents of fusants were slightly increased compared with wild types. Ethanol production by intraspecific and/or interspecific fusants was not increased, however, acetic acid production as byproduct was decreased or not detected, which indicates that industrial thermophilic anaerobes can be improved by means of protoplast fusion of two strains.