• The Plant Pathology Journal
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• 제29권4호
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• pp.460-464
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• 2013

dsRNA was found in malformed cultures of Lentinula edodes strain FMRI0339, one of the three most popular sawdust cultivated commercial strains of shiitake, and was also found in healthy-looking fruiting bodies and actively growing mycelia. Cloning of the partial genome of the dsRNA revealed the presence of the RdRp sequence of a novel L. edodes mycovirus (LeV), and sequence comparison of the cloned amplicon showed identical sequences sequence to known RNA-dependent RNA polymerase genes of LeV found in strain HKA. The meiotic stability of dsRNA was examined by measuring the ratio of the presence of dsRNA among sexual monokaryotic progeny. More than 40% of the monokaryotic progeny still contained the dsRNA, indicating the persistence of dsRNA during sexual reproduction. Comparing the mycelia growth of monokaryotic progeny suggested that there appeared to be a tendency toward a lower frequency of virus incidence in actively growing progeny.

• 한국버섯학회지
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• 제10권3호
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• pp.101-108
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• 2012

Pleurotus ostreatus 'Miso' is a mutant strain showing white color in pileus from the known parent strain 'Wonhyeong 1'. Shape and several other characters also vary with culture conditions. Mating experiments were performed to understand interstrain mating relationship using monokaryons of the parent and the mutant strains. All monokaryons were grown from single spores isolated from freshly collected fruit bodies. Pairings were performed in 90 mm petri dishes on PDA. They were allowed to grow at 25 until two fronts of the advancing mycelia met and developed a conspicuous contact zone. The contact zone and the outer edges of paired colonies on each plate were examined for clamp connections. The parent and the mutant resulted in tetrapolar incompatibility in intrastrain crosses. In interstrain crosses, each monokaryotic tester strain of the parent strain was out-crossed to monokaryotic tester strains of the mutant. As a result of these crosses it was found that both strains share the same A and B incompatibility factors yielding 25% compatibility.

• Mycobiology
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• 제45권4호
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• pp.379-384
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• 2017

In mating of Lentinula edodes, dikaryotic strains generated from certain monokaryotic strains such as the B2 used in this study tend to show better quality of fruiting bodies regardless of the mated monokaryotic strains. Unlike B2, dikaryotic strains generated from B16 generally show low yields, with deformed or underdeveloped fruiting bodies. This indicates that the two nuclei in the cytoplasm do not contribute equally to the physiology of dikaryotic L. edodes, suggesting an expression bias in the allelic genes of the two nuclei. To understand the role of each nucleus in dikaryotic strains, we investigated single nucleotide polymorphisms (SNPs) in laccase genes of monokaryotic strains to reveal nuclear origin of the expressed mRNAs in dikaryotic strain. We performed reverse transcription PCR (RT-PCR) analysis using total RNAs extracted from dikaryotic strains (A5B2, A18B2, and A2B16) as well as from compatible monokaryotic strains (A5, A18, and B2 for A5B2 and A18B2; A2 and B16 for A2B16). RT-PCR results revealed that Lcc1, Lcc2, Lcc4, Lcc7, and Lcc10 were the mainly expressed laccase genes in the L. edodes genome. To determine the nuclear origin of these laccase genes, the genomic DNA sequences in monokaryotic strains were analyzed, thereby revealing five SNPs in Lcc4 and two in Lcc7. Subsequent sequence analysis of laccase mRNAs expressed in dikaryotic strains revealed that these were almost exclusively expressed from B2-originated nuclei in A5B2 and A18B2 whereas B16 nucleus did not contribute to laccase expression in A2B16 strain. This suggests that B2 nucleus dominates the expression of allelic genes, thereby governing the physiology of dikaryons.

• 한국균학회소식:학술대회논문집
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• 한국균학회 2015년도 추계학술대회 및 정기총회
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• pp.37-37
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• 2015

dsRNA was found in malformed cultures of Lentinula edodes strain FMRI0339, one of the three most popular sawdust cultivated commercial strains of shiitake, and was also found in healthy-looking fruiting bodies and actively growing mycelia. Cloning of the partial genome of the dsRNA revealed the presence of the RdRp sequence of a novel L. edodes mycovirus (LeV), and sequence comparison of the cloned amplicon showed an identical sequence to known RdRp genes of LeV found in strain HKA. The meiotic stability of dsRNA was examined by measuring the ratio of the presence of dsRNA among sexual monokaryotic progeny. More than 40% of the monokaryotic progeny still contained the dsRNA, indicating the persistence of dsRNA during sexual reproduction. Comparing the mycelia growth of monokaryotic progeny suggested that, although variations in the growth rate existed among progeny and virus infection was observed in highly actively growing progeny, there appeared to be a tendency toward a lower frequency of virus incidence in actively growing progeny. This study attempted to cure the edible mushroom L. edodes strain FMRI0339 of the L. edodes mycovirus (LeV) in order to obtain an isogenic virus-free fungal strain as well as a virus-infected strain for comparison. Mycelial fragmentation, followed by being spread on a plate with serial dilutions resulted in a virus-free colony. Viral absence was confirmed with gel electrophoresis after dsRNA-specific virus purification, Northern blot analysis, and PCR using reverse transcriptase (RT-PCR). Once cured, all of fungal cultures remained virus-free over the next two years. Interestingly, the viral titer of LeV varied depending on the culture condition. The titer from the plate culture showed at least a 20-fold higher concentration than that grown in the liquid culture. However, the reduced virus titer in the liquid culture was recovered by transferring the mycelia to a plate containing the same medium. In addition, oxygen-depleted culture conditions resulted in a significant decrease of viral concentration, but not to the extent seen in the submerged liquid culture. Although no $discernable phenotypic changes in colony morphology were observed, virus-cured strains showed significantly higher growth rates and mycelial mass than virus-infected strains. We were also explored effects of LeV on fruiting body formation and mushroom yield. The fruiting body formation yield of virus-free L. edodes was larger than virus-infected L. edodes. These results indicate that LeV infection has a deleterious effect on mycelial growth and fruiting body formation. In addition, we have been investigated host-parasite interaction between L. edodes and its mycovirus interaction to study viral mechanism by establishment of proteomics.

• Mycobiology
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• 제43권1호
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• pp.81-86
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• 2015

To promote the selection of promising monokaryotic strains of button mushroom (Agaricus bisporus) during breeding, 61 progeny strains derived from basidiospores of two different lines of dikaryotic parental strains, ASI1038 and ASI1346, were analyzed by nucleotide sequencing of the intergenic spacer I (IGS I) region in their rDNA and by extracellular enzyme assays. Nineteen different sizes of IGS I, which ranged from 1,301 to 1,348 bp, were present among twenty ASI1346-derived progeny strains, while 15 different sizes of IGS I, which ranged from 700 to 1,347 bp, were present among twenty ASI1038-derived progeny strains. Phylogenetic analysis of the IGS sequences revealed that different clades were present in both the ASI10388- and ASI1346-derived progeny strains. Plating assays of seven kinds of extracellular enzymes (${\beta}$-glucosidase, avicelase, CM-cellulase, amylase, pectinase, xylanase, and protease) also revealed apparent variation in the ability to produce extracellular enzymes among the 40 tested progeny strains from both parental A. bisporus strains. Overall, this study demonstrates that characterization of IGS I regions and extracellular enzymes is useful for the assessment of the substrate-degrading ability and heterogenicity of A. bisporus monokaryotic strains.

• Mycobiology
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• 제49권1호
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• pp.86-88
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• 2021

The monokaryotic strain, Schizophyllum commune strain IUM1114-SS01, was generated from a basidiospore of dikaryotic parental strain IUM1114. It even showed the decolorizing activities for several textile dyes much better than its parental strain. Based on the results of a single-molecule real-time sequencing technology, we present the draft genome of S. commune IUM1114-SS01, comprising 41.1 Mb with GC contents of the genome were 57.44%. Among 13,380 protein-coding genes, 534 genes are carbon hydrate-active enzyme coding genes.

• Mycobiology
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• 제46권4호
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• pp.407-415
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• 2018

Pheromone (PHB)-receptor (RCB) interaction in the mating pheromone response pathway of Lentinula edodes was investigated using synthetic PHBs. Functionality of the C-terminally carboxymethylated synthetic PHBs was demonstrated by concentration-dependent induction of a mating-related gene (znf2) expression and by pseudoclamp formation in a monokaryotic strain S1-11 of L. edodes. Treatment with synthetic PHBs activated the expression of homeodomain genes (HDs) residing in the A mating type locus, and of A-regulated genes, including znf2, clp1, and priA, as well as genes in the B mating type locus, including pheromone (phb) and receptor (rcb) genes. The synthetic PHBs failed to discriminate self from non-self RCBs. PHBs of the B4 mating type (B4 PHBs) were able to activate the mating pheromone response pathway in both monokaryotic S1-11 and S1-13 strains, whose B mating types were B4 (self) and B12 (non-self), respectively. The same was true for B12 PHBs in the B4 (non-self) and B12 (self) mating types. The synthetic PHBs also promoted the mating of two monokaryotic strains carrying B4-common incompatible mating types ($A5B4{\times}A1B4$). However, the dikaryon generated by this process exhibited abnormally high content of hyphal branching and frequent clamp connections and, more importantly, was found to be genetically unstable due to overexpression of mating-related genes such as clp1. Although synthetic PHBs were unable to discriminate self from non-self RCBs, they showed a higher affinity for non-self RCBs, through which the mating pheromone response pathway in non-self cells may be preferentially activated.

• 한국균학회지
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• 제49권4호
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• pp.487-495
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• 2021

표고는 사극성의 교배계를 갖는 담자균의 일종으로, 표고의 교배형은 A와 B라 불리는 서로 독립된 두 유전자좌에 의해 결정된다. 이론적으로 하나의 이핵균주는 네 개의 서로 다른 교배형을 갖는 담자포자를 1:1:1:1의 비율로 만들 수 있다. 과거 연구 결과에 따르면, 표고 담자포자에서 교배형이 편향된 분리비로 나타남이 보고되었다. 하지만 이러한 결과들은 꺽쇠연결과 같은 형태학적 특성만을 기반으로 교배형을 결정했다는 한계가 있다. 이에 본 연구에서는 교배형의 편향된 분리비가 표고에서 일반적인 현상인지 보다 명확하게 알아보기 위해서 최근에 보고된 DNA 마커를 활용하여 세 가지 표고 품종들의 담자포자에 대한 교배형 분석을 수행하였다. 그 결과 교배형의 편향된 분리비가 과거 보고와 일치하게 균주 특이적인 특성임을 확인하였다. 분석한 세 품종 중 한 품종을 제외하고 나머지 두 품종에서 편향된 분리비가 관찰된 것이다. 다음으로는 각 담자포자 유래 단핵균사들의 생장 속도와 교배형의 상관관계를 분석하였다. 그 결과 표고 단핵균사의 생장 속도는 B 교배형과는 관계가 없고, A 교배형과 관계가 있음을 확인하였다. 따라서 A 교배형 유전자좌에 존재하는 호메오도메인 전사인자 혹은 A 교배형 유전자좌와 연관된 유전자들이 단핵균사의 생장에 영향을 줄 것으로 보인다. 버섯 신품종 육성에서 교배형의 중요성을 고려할 때, 본 연구는 효율적인 신품종 육성 전략을 세우거나 단핵균사 생장 기작을 이해하는 데 도움을 줄 것으로 기대된다.

• 한국균학회지
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• 제50권4호
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• pp.263-274
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• 2022

표고는 중요한 상업용 버섯이며, 표고에서의 바이러스 감염에 대한 여러 보고가 있었다. 톱밥 재배 품종인 산백향(NIFoS 2778)과 태향고(NIFoS 4317)에 대한 바이러스 검출 결과, 이들이 2개의 mycoviruses (LeV-HKB 및 LeNSRV1)에 감염되어 있음을 확인하였다. 각 품종의 자실체에서 분리된 담자포자에서 유래한 80개의 단핵균사에서 바이러스 감염을 조사한 결과, 대부분의 단 핵균사들이 바이러스에 감염되어 있었으며, LeV-HKB와 LeNSRV1의 수직감염률에 차이가 있었다. LeV-HKB가 LeNSRV1 보다 높은 수직감염률을 나타낸 것이다. 따라서 mycovirus의 수직 감염 기작이 바이러스 종에 따라 다른 것으로 보인다. 다음으로, 담자포자 유래 단핵균사들의 생장속도를 조사하여 바이러스 감염과 생장속도의 상관관계를 조사하였다. 바이러스 감염과 균사 생장속도 사이에 통계적으로 유의한 상관관계가 없었지만, 감염된 바이러스의 종류가 늘어날수록 생장속도가 감소하는 경향을 확인하였다. 본 연구는 mycovirus의 수직 감염 기작을 이해하는 데 기여하고, 바이러스에 감염되지 않은 단핵균사를 이용한 무바이러스 품종 개발을 촉진하는 데 기여할 수 있을 있을 것 기대된다.

• 한국균학회소식:학술대회논문집
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• 한국균학회 2014년도 추계학술대회 및 정기총회
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• pp.42-42
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• 2014

Mating of tetrapolar mushrooms is regulated by to chromosomal loci, A and B. A locus contains A gene that expresses a homeodomain protein whereas B locus contains multiple pheromones and receptor genes. In order to characterize the mating loci in Korean cultivated strains of Lentinula edodes, one hundred monokaryotic myclelia were isolated from the basidiospores of cultivated strains, including Cham-A-Ram, Sanjo701, and Sanjo707. Both mating loci were amplified using primer sets targeting conserved sequence regions for homeodomain (HD), pheromone, and receptor genes. Subsequent sequence analysis revealed that the Korean strains contained significant variations in the homeodomain of A locus, even within the same A1 or A2 mating type. Similarly, B locus was also highly diversified in the sequences of pheromones and receptors as well as gene organization. These results enabled us to design mating type-specific probes which can distinguish mating type of each strain. The specificity was confirmed by between intra- and inter-strain mating experiment.