Background: Following years of inactivity, psychedelic research has rapidly expanded within clinical and therapeutic fields. In particular, magic mushrooms (psilocybin), a plant-based psychedelic, have been researched for the treatment of complex mental health and substance dependence conditions, and yielded promising results. Largely due to the historical baggage of the psychedelic movement in the 1950s-1970s, and the stigma of recreational substance use, recreational magic mushroom users have been ignored within the current psychedelic revival. This thesis addressed this gap, examining the magic mushroom recreational substance use patterns of emerging adults in Victoria, British Columbia. Theory and Methods: Using the normalization thesis as the guiding theoretical framework, this thesis used a sequential-exploratory mixed methods design. Statistical analysis of quantitative cross-sectional interviews (n=558) conducted between 2008 -2016 generated rates of use, availability, and self-rated knowledge rates of magic mushrooms users. Qualitative cross-sectional semi-structured interviews (n=20) analyzed through thematic analysis determined substance use behaviors with reference to the current social and cultural context. Participants were recreational magic mushroom users, aged 19- 24. Results: Quantitative results indicated high overall rates of lifetime and past year magic mushroom use, with the lowest reported prevalence rate of lifetime use occurring in 2014 (86%), suggesting high rates of use within the recreational substance using population. There were no statistically significant relationships between year and lifetime or past year rates magic mushroom use. Gender was statistically significantly associated with magic mushroom use, with males being more likely to use magic mushrooms. Qualitative results indicated dynamic and strategically planned magic mushroom experiences. Themes developed include: shifting understandings, optimizing experience, purpose driven use; and post-trip impact. Participants reported using for transformational, micro-dosing, and leisure purposes. Discussion: Results suggested that magic mushroom use is in the process of differentiated normalization and assimilative normalization, influenced by developmental, social and cultural forces. Recreational users report substance use practices that have not been widely reported with the substance use literature, including using small doses of magic mushrooms (i.e. micro-dosing) for self-enhancement and therapeutic purposes. Results can be applied to the current psychedelic revival in three ways: (1) directing future clinical research directions and; (2) provide lived and experience and relevancy to clinical research, which will improve applicability and; (3) re-conceptualizing the identity of a recreational substance user, which has important implications regarding stigmatization. / Graduate

Extracts from medicinal mushrooms were prepared and tested for anti-microbial activity against food pathogens and food spoilage microorganisms. The inhibitory activity was measured using a disk diffusion assay and with optical density (OD). For OD, 7 fractions were collected using HPLC for 4 (A. blazei Murrill, G. lucidum, G. frondosa, I. obiquus) medicinal mushrooms and 6 fractions from L. edodes and 8 fractions from P. linteus. The results from disk diffusion assay showed that most mushrooms displayed significant inhibition compared to the ethanol. The exceptions were: A. blazei Murrill, I. obliquus, and L. edodes against E. coli O157:H7; I. obliquus against L. monocytogenes V7; I. obliquus against S. cerevisiae Y99; L. edodes against Z. bailii Y03; and I. obliquus against Z. bailii/bisporus Y108. Inhibition was more effective in yeasts than bacteria. The result from Bioscreen C showed that against L. monocytogenes V7, fraction 7 in A. blazei Murrill; fraction 1, 4 and 5 in G. lucidum; fraction 4 in G. frondosa; and fraction 4 and 5 in I. obliquus significantly inhibited the growth compared to ethanol. Against Z. bailii Y03, fraction 7 in A. blazei Murrill; all fractions from G. lucidum, G. frondosa, and P. linteus; fraction 1, 2, 3, and 6 from I. obliquus; and fraction 4 and 6 from L. edodes significantly inhibited growth compare to ethanol. The results indicated that there is significant antimicrobial activity against food pathogens and spoilage organisms in the medicinal mushrooms studied. / Ph. D.

Orientador: Marlene Rita de Queiroz / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Engenharia Agrícola / Made available in DSpace on 2018-08-17T11:09:06Z (GMT). No. of bitstreams: 1 Sampaio_SaraMedeiros_M.pdf: 362284 bytes, checksum: 9f40de09eb4c685cbda1a3c211ebaf16 (MD5) Previous issue date: 2003 / Resumo: A finalidade principal deste trabalho foi reunir subsídios para orientar os processos desecagem, armazenagem e reidratarão do cogumelo Shiitake. A pesquisa foi realizada durante o ano de 2002 no Laboratório de Tecnologia Pós-Colheita/Secagem da Faculdade de Engenharia Agrícola da Universidade Estadual de Campinas e consistiu em duas etapas: secagem e armazenagem por 3 meses. Os fatores estudados foram: na secagem - geometria de corte (Shiitake inteiro e fatiado), temperatura de secagem (50ºC e 70°C), teor de umidade final (5% e 15%); na armazenagem - geometria de corte (Shiitake inteiro e fatiado), teor de umidade inicial (5% e 15%), embalagem (com e sem saco de polipropileno) e tempo de armazenagem (3 meses); na reidratação - tempo de imersão em água (8 tempos), geometria de corte (Shiitake inteiro e fatiado), teor de umidade inicial (5% e 15%) e tempo de armazenagem (3 meses). Os parâmetros de qualidade estudados foram: cor, textura, massa, teor de umidade, capacidade de reidratação. Os parâmetros de secagem estudados foram: cinéticas experimentais de secagem. As cinéticas de secagem mostraram que a secagem ocorreu no período de taxa decrescente e que a secagem dos cogumelos fatiados secos à 70ºC ocorreu em menor tempo que os demais tratamentos. A temperatura de secagem de 70ºC proporcionou menor escurecimento e os resultados do efeito da secagem sobre a textura do cogumelo foram pouco conclusivos. O tempo de armazenagem afetou a qualidade dos cogumelos ocorrendo maior dureza, gomosidade e escurecimento. Entretanto, o uso da embalagem retardou o escurecimento. Cogumelos fatiados armazenados com 15% de umidade inicial e embalados obtiveram pouca variação de mastigabilidade, teor de umidade e massa durante a armazenagem. O tempo de armazenagem provocou redução da capacidade de reidratação para os cogumelos com 5% de umidade inicial. Os cogumelos fatiados, com 5% de umidade inicial e embalados obtiveram valores de reidratação superiores aos demais tratamentos / Abstract: Mushroom (Shiitake), drying, storage and re-hydration: effect on quality parameter process determinations. To orient drying, storage and re-hydration processes, accumulated subsidies by studying some parameters on mushroom Shiitake. During three months, drying, storage and re-hydration processes, were studied on mushroom in 2002 at Drying/Post-harvesting Technology Lab from Agricultural Engineering School of the Campinas State University. In drying process were studied: drying temperature (50º and 70 ºC), final moisture content (5% and 15%), cutting geometry (mushroom whole and sliced); in re-hydration process besides drying temperature and cutting geometry, were assessed also, water immersion (8 times), initial moisture content (5%% and 15%) and storage time (3 months); in packing up process; (with/without polypropylene bag), storage time (3 months) and initial moisture content were evaluated. Color, texture, mass, moisture content, and re-hydration capacity, were also evaluated. Drying kinetics showed that drying occurred at increased rate in the period and sliced mushroom dried at 70º C, occurred in lesser time than other treatments. Drying temperature at 70 ºC had less darkening and the results on mushroom texture drying effects were not conclusive. Drying time affected mushroom quality, occurring great hardness, gummosis and darkening. However, packing up use, delayed the darkening. Stored sliced and wrapped mushrooms with 15% initial moisture content had less, chew property variation, moisture content and mass during storage. Storage time provoked rehydration capacity decrease on mushrooms with 5% of initial moisture. Sliced and wrapped mushrooms, with 5% initial moisture, had re-hydration values greater than other treatments / Mestrado / Tecnologia Pós-Colheita / Mestre em Engenharia Agrícola

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by Shi Yuling. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2001. / Includes bibliographical references (p. 159-184). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. Ann Arbor, MI : ProQuest Information and Learning Company, [200-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Coprinopsis cinerea is extensively used as a model to study the development of homobasidiomycete fungi. Unraveling the molecular basis of fungal developmental processes would contribute to evolutionary studies, improve the knowledge about multicellularity development, and lead to improvement in the breeding and cultivation of edible or medical homobasidiomycete mushrooms. / I studied the fungal developmental processes in two aspects: 1) the hypothesis of a “developmental hourglass”, and 2) the existence of microRNA-like RNA (milRNA) genes in fungi. / The model of “developmental hourglass” suggests that middevelopment is the most conserved and the most resistant to evolutionary changes. Although extensively explored in animals and plants, such hourglass pattern has not been reported in fungi yet. I tested the hourglass model in C. cinerea using two complementary approaches, the transcriptome age index (TAI) and the transcriptome divergence index (TDI). Both the TAI and the TDI profiles displayed an hourglass pattern over the development of C. cinerea; the young fruiting body stage was the waist that expressed the evolutionarily oldest transcriptome (lowest TAI) and gave the strongest signal of purifying selection (lowest TDI). By cross-kingdom comparisons, it is found that all three kingdoms displayed high expression levels of genes in “information storage and processing” at the waist stage, while genes in “metabolism” became more active later; besides, genes at the waist stage were underrepresented in “signal transduction mechanisms”. / MicroRNA (miRNA) is a group of endogenous non-coding regulatory RNAs of ~22 nt that regulate gene expression in various biological processes such as cell differentiation, development regulation and heterochromatin formation. Past research work reported several simple filamentous fungi to contain milRNAs in their genomes; however, no milRNA has been reported in mushrooms so far. Through computational prediction, I identified 16 putative milRNA genes in C. cinerea. Besides, evolutionary analysis showed that C. cinerea contains Dicer-like proteins (DCLs), which may play roles in milRNA biogenesis. / Both the discovery of a “developmental hourglass” and milRNA genes laid a foundation for analysis of fruiting body formation in fungi, and for evolutionary analysis of multicellular development across kingdoms. / 灰蓋鬼傘（Coprinopsis cinerea）作為一個典型的生物模型，被廣泛地運用在擔子菌生長發育的研究中。從分子層面上解析出真菌生長發育的過程，可以促進進化生物學的研究，提高對多細胞生物演化的認知，以及改善食用菌和藥用菌的育種與培養。 / 我從兩個角度去分析了真菌的生長發育過程：1）真菌的“發育沙漏”假說，2）真菌基因組裡類微RNA（microRNA-like RNA [milRNA]）的基因。 / “發育沙漏”模型指出，發育中期在進化中是最保守的、最能抵抗進化帶來的改變。這個沙漏模型在動物與植物中被廣泛地研究與證實，但是迄今為止，並未有人對其在真菌中的存在進行過探究。我以C. cinerea作為研究模型，採用了兩種互補的方法，transcriptome age index (TAI) 和 transcriptome divergence index (TDI)，探究了發育沙漏在真菌中的存在。兩個指數均在C. cinerea發育過程中呈現出了沙漏的形狀；年輕的子實體（young fruiting body）階段是沙漏的腰身，表達出進化中最古老的基因並給出了最強的淨化選擇（purifying selection）的信號。我將三大真核生物進行了比較，發現在腰身階段時，在“信息儲存和處理”中發揮作用的基因呈現出了高表達量，而“代謝”基因的表達量在後期更高；而且在腰身階段表達的基因中，負責“信號傳遞機制”的基因偏少。 / 微RNA（microRNA [miRNA]）是一類長約22個核苷酸、不轉譯蛋白質的RNA分子。它們可以調節其他基因的表達，在多方面的生理過程中發生作用，比如細胞分化、發育調節、異染色質的形成等等。近幾年的研究表明，一些簡單的絲狀真菌的基因組中含有類微RNA（milRNA）的基因；但是，至今未有報導說蘑菇是否含有此類基因。我運用計算預測的方法在C. cinerea的基因組裡找到了16個可能的類微RNA基因。此外，從進化的角度分析，我發現C. cinerea含有類Dicer蛋白酶（Dicer-like proteins [DCLs]），而這些類Dicer蛋白酶可能在milRNA的產生過程中發揮作用。 / 這篇論文所報導的兩個發現──真菌的“發育漏斗“和類微RNA基因，均為之後子實體發育的研究及多細胞發育的研究奠定了基礎。 / Cheng, Xuanjin. / Thesis Ph.D. Chinese University of Hong Kong 2015. / Includes bibliographical references. / Abstracts also in Chinese. / Title from PDF title page (viewed on 09, September, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.

by To Siu-wing. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves 182-195). / ACKNOWLEDGEMENTS --- p.VI / ABSTRACT --- p.VII / LIST OF TABLES --- p.IX / LIST OF FIGURES --- p.XI / ABBREVIATIONS --- p.XVII / Chapter PART I --- GENERAL ASPECTS / Chapter CHAPTER 1 --- GENERAL INTRODUCTION --- p.1 / Chapter CHAPTER 2 --- LITERATURE REVIEW / Chapter 2.1. --- History of fungal protoplast fusion / Chapter 2.1.1. --- Fungal protoplast preparation technique --- p.4 / Chapter 2.1.2. --- Application of fungal protoplasts --- p.5 / Chapter 2.2. --- Protoplast fusion by polyethene glycol (PEG) --- p.9 / Chapter 2.3. --- Incompatibility system in fungi --- p.10 / Chapter 2.4. --- Characterization of fusion products by genetic markers --- p.12 / Chapter PART II --- OPTIMIZATION OF PROTOPLAST RELEASE AND PROTOPLAST FUSION STUDIES / Chapter CHAPTER 3 --- PROTOPLAST ISOLATION OF Pleurotus florida AND Schizophyllum commune / Chapter 3.1. --- Introduction --- p.14 / Chapter 3.2. --- Materials and methods / Chapter 3.2.1. --- Strains and culture media --- p.14 / Chapter 3.2.2. --- Protoplast isolation in different types and concentrations of lytic enzyme --- p.15 / Chapter 3.2.3. --- Protoplast isolation using mycelium with different culture ages --- p.17 / Chapter 3.2.4. --- Protoplast isolation in different types and concentrations of osmotic stabilizers --- p.17 / Chapter 3.2.5. --- Collection of protoplast by centrifugation --- p.18 / Chapter 3.3. --- Results / Chapter 3.3.1. --- Effect of type and concentration of lytic enzyme --- p.19 / Chapter 3.3.2. --- Efficiency of protoplast isolation from mycelia with different culture ages --- p.25 / Chapter 3.3.3. --- Effect of types and concentrations of osmotic stabilizers --- p.28 / Chapter 3.3.4. --- Collecting efficiency of protoplast by centrifugation --- p.31 / Chapter 3.4. --- Discussion / Chapter 3.4.1. --- Choice of lytic enzyme system and time for enzyme digestion --- p.33 / Chapter 3.4.2. --- Culture age for maximum protoplast yield --- p.34 / Chapter 3.4.3. --- Choice of concentration and type of osmotic stabilizers --- p.35 / Chapter CHAPTER 4 --- PROTOPLAST FUSION OF Pleurotus florida AND Schizophyllum commune / Chapter 4.1. --- Introduction --- p.38 / Chapter 4.2. --- Materials and methods / Chapter 4.2.1. --- Protoplast formation and size of protoplasts --- p.39 / Chapter 4.2.2. --- Fluorescent staining of protoplasts' nuclei --- p.39 / Chapter 4.2.3. --- Stability of the genetics markers / Chapter 4.2.3.1. --- Preparation of media for checking the presence of genetics markers --- p.40 / Chapter 4.2.3.2. --- Determining the presence of auxotrophic as well as drug resistance markers --- p.42 / Chapter 4.2.4. --- Regeneration of mycelium from protoplast --- p.42 / Chapter 4.2.5. --- Protoplast fusion and screening of fusion products --- p.45 / Chapter 4.3. --- Results / Chapter 4.3.1. --- Size of protoplasts ofPf67 and Scl7 --- p.48 / Chapter 4.3.2. --- Proportion of protoplasts bearing nucleus --- p.48 / Chapter 4.3.3. --- Protoplast regeneration in regeneration medium / Chapter 4.3.3.1. --- Protoplasts regeneration morphologies --- p.52 / Chapter 4.3.3.2. --- Regeneration frequencies and back mutation frequencies of Pf67 and Scl7 protoplasts --- p.58 / Chapter 4.3.4. --- Effect of PEG fusion treatment on auxotrophic and drug resistance markers of Pf67 and Scl7 --- p.60 / Chapter 4.3.5. --- Fusion products obtained from screening process --- p.61 / Chapter 4.4. --- Discussion / Chapter 4.4.1. --- Effect of protoplast isolation and PEG treatment on the two fusion parents --- p.63 / Chapter 4.4.2. --- Structural heterogeneity of protoplasts --- p.64 / Chapter 4.4.3. --- Polymorphic nature of protoplast regeneration --- p.67 / Chapter 4.4.4. --- Protoplast fusion frequence --- p.67 / Chapter PART III --- ANALYSIS OF FUSION PARENTS AND FUSION PRODUCTS / Chapter CHAPTER 5 --- MORPHOLOGICAL AND CYTOLOGICAL STUDIES / Chapter 5.1. --- Introduction --- p.69 / Chapter 5.2. --- Materials and methods / Chapter 5.2.1. --- Strains --- p.69 / Chapter 5.2.2. --- Study on colonial and mycelial morphology --- p.70 / Chapter 5.2.3. --- Fluorescent staining of mycelial nuclei with DAPI --- p.70 / Chapter 5.2.4. --- Study on fruit body and basidial morphology / Chapter 5.2.4.1. --- Fruiting on agar plate --- p.71 / Chapter 5.2.4.2. --- Scanning electron microscopic examination --- p.73 / Chapter 5.3. --- Results / Chapter 5.3.1. --- Variation of colonial morphology --- p.74 / Chapter 5.3.2. --- Morphologies and the number of nuclei in the mycelial cells of fusion parents and fusion products --- p.76 / Chapter 5.3.3. --- Fruit body morphology --- p.82 / Chapter 5.3.4. --- Basidial morphology --- p.84 / Chapter 5.4. --- Discussion --- p.87 / Chapter CHAPTER 6 --- PHYSIOLOGICAL STUDIES OF FUSION PARENTS AS WELL AS FUSION PRODUCTS BY INVESTIGATING THE GROWTH RESPONSES TO DRUGS / Chapter 6.1. --- Introduction --- p.90 / Chapter 6.2. --- Materials and methods / Chapter 6.2.1. --- Strains and media --- p.96 / Chapter 6.2.2. --- Growth responses of the strains to different concentrations of drugs --- p.97 / Chapter 6.3. --- Results / Chapter 6.3.1. --- Comparison of growth pattern as well as growth rate between fusion parents and fusion regenerants --- p.98 / Chapter 6.3.2. --- Growth responses of fusion parents and fusion products on complete medium --- p.105 / Chapter 6.3.3. --- Growth responses of fusion parents and fusion regenerants on complete medium with acriflavin --- p.108 / Chapter 6.3.4. --- Growth responses of fusion parents and fusion products on complete medium with guaiacol --- p.111 / Chapter 6.4. --- Discussion / Chapter 6.4.1. --- General considerations on experimental design --- p.115 / Chapter 6.4.2. --- Growth responses of protoplast regenerants of either fusion parents --- p.116 / Chapter 6.4.3. --- Growth responses on complete medium without fungitoxic drug --- p.117 / Chapter 6.4.4. --- Growth responses on the acriflavin agar medium --- p.118 / Chapter 6.4.5. --- Growth responses on guaiacol agar medium --- p.119 / Chapter 6.4.6. --- Summary --- p.120 / Chapter CHAPTER 7 --- GENETICAL STUDIES / Chapter 7.1. --- Introduction --- p.121 / Chapter 7.2. --- Materials and methods / Chapter 7.2.1. --- Segregation tests of auxotrophic and drug resistance markers in progeny of dikaryotic fusion product --- p.127 / Chapter 7.2.2. --- Complementation test of fusion products as well as the spore germinants of dikaryotic fusion product PS1 --- p.129 / Chapter 7.2.3. --- Recovery of the individual nuclear type of dikaryotic fusion product PS1 --- p.130 / Chapter 7.2.4. --- Genomic fingerprinting / Chapter 7.2.4.1. --- Strains and culture medium --- p.133 / Chapter 7.2.4.2. --- Genomic DNA preparation by cesium chloride (CsCl) method --- p.135 / Chapter 7.2.4.3. --- Genomic DNA preparation by chloroform :TE saturated phenol method --- p.136 / Chapter 7.2.4.4. --- Qualitative analysis of genomic DNA --- p.137 / Chapter 7.2.4.5. --- Quantitative analysis of genomic DNA --- p.137 / Chapter 7.2.4.6. --- DNA amplification by arbitrarily primed -polymerase chain reaction --- p.138 / Chapter 7.3. --- Results / Chapter 7.3.1. --- Progeny analysis and determination of auxotrophic as well as drug resistance markers --- p.140 / Chapter 7.3.2. --- Complementation tests of the fusion products as well as the spore germinants of dikaryotic fusion product PS1 --- p.143 / Chapter 7.3.3. --- Monokaryotic protoplast regenerants of dikaryotic fusion product PS1 --- p.147 / Chapter 7.3.4. --- Studies on extraction of undigested genomic DNA --- p.148 / Chapter 7.3.5. --- Genomic fingerprinting by AP-PCR --- p.155 / Chapter 7.4. --- Discussion / Chapter 7.4.1. --- Genomic DNA extraction --- p.161 / Chapter 7.4.2. --- Recovery of the individual nuclear type of dikaryotic fusion product PS1 --- p.165 / Chapter 7.4.3. --- Genomic changes in fusion products --- p.167 / Chapter 7.4.4. --- Progeny analysis and determination of auxotrophic as well as drug resistance markers --- p.171 / Chapter PART IV --- SUMMING-UP / Chapter CHAPTER 8 --- GENERAL SUMMARY AND CONCLUSION REMARKS / Chapter 8.1. --- General summary --- p.176 / Chapter 8.2. --- Conclusion remarks and future studies --- p.179 / REFERENCES --- p.182 / APPENDIX A SOLUTIONS

by Jiong Zhao. / Thesis (Ph.D.)--Chinese University of Hong Kong, 1994. / Includes bibliographical references (leaves 197-217). / Acknowledgments --- p.III / Abstract --- p.IX / Abbreviations --- p.XI / Chapter Chapter 1. --- General Introduction --- p.1 / Chapter 1.1 --- What is a mushroom? --- p.1 / Chapter 1.2 --- Mushroom Genetics: its development and prospective --- p.1 / Chapter 1.2.1 --- Genome karyotype by pulsed field gel electrophoresis analysis --- p.2 / Chapter 1.2.2 --- Mitochondrial Genetics --- p.4 / Chapter 1.2.3 --- Mating type genes --- p.5 / Chapter 1.2.4 --- Transformation --- p.7 / Chapter 1.2.5 --- Parasexual processes --- p.8 / Chapter 1.2.6 --- Mushroom breeding --- p.11 / Chapter Chapter 2. --- Literature review: Protoplast fusion in fungi --- p.14 / Chapter 2.1 --- Introduction --- p.14 / Chapter 2.2 --- Protoplast fusion in yeasts --- p.14 / Chapter 2.2.1 --- Intraspecific fusion --- p.14 / Chapter 2.2.2 --- Interspecific fusion --- p.15 / Chapter 2.2.3 --- Intergeneric fusion --- p.16 / Chapter 2.3 --- Protoplast fusion in some Filamentous fungi --- p.17 / Chapter 2.3.1 --- Aspergillus --- p.17 / Chapter 2.3.2 --- Fusarium --- p.18 / Chapter 2.3.3 --- Tricoderma --- p.19 / Chapter 2.4 --- Protoplast fusion in strains --- p.21 / Chapter 2.4.1 --- Protoplast isolation and regeneration --- p.21 / Chapter 2.4.2 --- Intraspecific fusion in mushroom species --- p.24 / Chapter 2.4.3 --- Interspecific fusion in mushroom species --- p.24 / Chapter 2.4.4 --- Intergeneric fusion in mushroom species --- p.26 / Chapter 2.4.5 --- Transfer of nuclei in mushroom species --- p.27 / Chapter 2.5 --- General conclusions about literatures --- p.27 / Chapter 2.5.1 --- Brief points about fungal protoplast fusion --- p.27 / Chapter 2.5.2 --- Some arguements about fusion works in mushrooms strains --- p.31 / Chapter 2.5.2.1 --- Classification of parental strains --- p.31 / Chapter 2.5.2.2 --- Control experiments --- p.31 / Chapter 2.5.2.3 --- Indentification methods of hybrids --- p.32 / Chapter 2.6 --- General research ideas about experiments --- p.33 / Chapter Chapter 3 --- Protoplast isolation and regeneration in some mushroom species --- p.37 / Chapter 3.1 --- Introduction --- p.37 / Chapter 3.2 --- Materials and Methods --- p.38 / Chapter 3.2.1 --- Strains --- p.38 / Chapter 3.2.2 --- Media --- p.38 / Chapter 3.2.3 --- Protoplast release --- p.40 / Chapter 3.2.4 --- Protoplast regeneration --- p.41 / Chapter 3.3 --- Results and Discussion --- p.41 / Chapter 3.3.1 --- Effect of culture age --- p.41 / Chapter 3.3.2 --- Effect of lytic enzyme --- p.42 / Chapter 3.3.3 --- Effect of concentration of mycelium --- p.45 / Chapter 3.3.4 --- Effect of filter system --- p.46 / Chapter 3.3.5 --- Effect of different regeneration protocols --- p.48 / Chapter 3.3.6 --- Effect of soluable starch --- p.49 / Chapter 3.3.7 --- Effect of PEG on the regeneration frequency --- p.50 / Chapter 3.4 --- Conclusions --- p.53 / Chapter Chapter 4 --- Monokaryotization by protoplasting technique in some heterothallic mushroom species --- p.54 / Chapter 4.1 --- Introduction --- p.54 / Chapter 4.2 --- Materials and Methods --- p.55 / Chapter 4.2.1 --- Strains and media --- p.55 / Chapter 4.2.2 --- Production of neo-monokaryons by protoplast technique --- p.55 / Chapter 4.2.3 --- Identification of mating types in protoplasted monokaryons --- p.57 / Chapter 4.3 --- Results / Chapter 4.3.1 --- Formation of neo-monokaryons --- p.57 / Chapter 4.3.2 --- Monokaryotization in different strains --- p.60 / Chapter 4.3.3 --- Comparison of parental and protoplasted monokaryons --- p.60 / Chapter 4.3.4 --- Comparison of regeneration rate of parental monokaryons --- p.62 / Chapter 4.4 --- Discussion / Chapter 4.4.1 --- Differences of regeneration time in monokaryons and dikaryons --- p.64 / Chapter 4.4.2 --- Genetic differences between parental and neo-monokaryons --- p.64 / Chapter 4.4.3 --- Mechanism for the production of neo-monokaryons --- p.65 / Chapter 4.4.4 --- Advantages of protoplasting technique in mushroom breeding --- p.65 / Chapter 4.4.5 --- Protoplasting technique in the identification of fusion hybrids --- p.67 / Chapter 4.5 --- Couclusions --- p.68 / Chapter Chapter 5 --- Intraspecific hybridization in Coprinus cinereus and Schizophyllum commune by PEG-induced protoplast fusion and electrofusion --- p.69 / Chapter 5.1 --- Introduction --- p.69 / Chapter 5.2 --- Materials and Methods / Chapter 5.2.1 --- Strains and Media --- p.70 / Chapter 5.2.2 --- Fusogen --- p.70 / Chapter 5.2.3 --- Inactivation chemicals --- p.71 / Chapter 5.2.4 --- Inactivation of protoplasts --- p.71 / Chapter 5.2.5 --- PEG induced protoplast fusion --- p.72 / Chapter 5.2.6 --- Electrofusion --- p.72 / Chapter 5.2.7 --- Investigation of protoplast fusion yield and fusion frequency --- p.73 / Chapter 5.2.8 --- Comparison of mycelium growth rate --- p.73 / Chapter 5.2.9 --- Fruiting test --- p.74 / Chapter 5.3 --- Results / Chapter 5.3.1 --- Inactivation by IA and DP --- p.76 / Chapter 5.3.2 --- Effect of different fusogens on fusion frequency --- p.79 / Chapter 5.3.3 --- Effect of different fusion protocols on fusion frequency --- p.79 / Chapter 5.3.4 --- Optimization of electrofusion --- p.80 / Chapter 5.3.5 --- Fusion frequency resulted by PEG and electrofusion --- p.83 / Chapter 5.3.6 --- Comparison of colony diameters and fruiting time --- p.84 / Chapter 5.4 --- Discussion / Chapter 5.4.1 --- Inactivation of protoplasts by biochemical inhibitors --- p.85 / Chapter 5.4.2 --- Optimization of PEG induced fusion --- p.86 / Chapter 5.4.3 --- Optimization of electrofusion --- p.86 / Chapter 5.4.4 --- Identification of fusion heterokaryons --- p.87 / Chapter 5.4.5 --- Comparison of PEG and electrofusion --- p.89 / Chapter 5.4.2 --- Effect of mitochondria --- p.90 / Chapter 5.5 --- Couclusions --- p.91 / Chapter Chapter 6 --- Interspecific hybridization between Volvariella volvacea and Volvariella bomhycina by protoplast fusion --- p.92 / Chapter 6.1 --- Introduction --- p.92 / Chapter 6.2 --- Materials and Methods / Chapter 6.2.1 --- Strains and Media --- p.93 / Chapter 6.2.2 --- Protoplast production and regeneration --- p.94 / Chapter 6.2.3 --- Inactivation of protoplasts --- p.94 / Chapter 6.2.4 --- Protoplast fusion --- p.94 / Chapter 6.2.5 --- Selection of fusion products --- p.95 / Chapter 6.2.6 --- Analyses of progeny --- p.95 / Chapter 6.2.7 --- Identification of fusants by protoplasting technique --- p.96 / Chapter 6.2.8 --- Nuclear DNA contents in parents and hybrids --- p.96 / Chapter 6.2.9 --- Genomic DNA amplification by arbitraly primers --- p.96 / Chapter 6.2.10 --- Amplification by nuclear and mitochondrial rDNA --- p.97 / Chapter 6.2.11 --- Fruiting test --- p.97 / Chapter 6.3 --- Results / Chapter 6.3.1 --- Inactivation of Vb10 protoplasts --- p.98 / Chapter 6.3.2 --- Low temperature effect on Vv34 --- p.100 / Chapter 6.3.3 --- Selection of fusants --- p.100 / Chapter 6.3.4 --- Analyses of progeny --- p.106 / Chapter 6.3.5 --- Identification by protoplasting technique --- p.108 / Chapter 6.3.6 --- Nuclear DNA contents in parents and hybrids --- p.110 / Chapter 6.3.7 --- Arbitraly primer amplified PCR fingerprinting --- p.113 / Chapter 6.3.8 --- rDNA PCR results --- p.119 / Chapter 6.3.9 --- Interspecific variations / Chapter 6.3.10 --- Genome analysis of hybrids by pulse field gel electrophoresis / Chapter 6.3.11 --- Fruiting test / Chapter 6.4 --- Discussion / Chapter 6.4.1 --- Strain choice --- p.125 / Chapter 6.4.2 --- Low temperature strains --- p.125 / Chapter 6.4.3 --- Nuclear DNA content --- p.125 / Chapter 6.4.4 --- AP-PCR and RAPDs markers --- p.126 / Chapter 6.4.5 --- Interspecific fusion in Volvariella --- p.126 / Chapter 6.5 --- Couclusions --- p.130 / Chapter Chapter 7 --- Intergeneric hybridization between Schizophyllum commune and Pleurotus florida by protoplast fusion --- p.131 / Chapter 7.1 --- Introduction --- p.131 / Chapter 7.2 --- Materials and Methods / Chapter 7.2.1 --- Strains and Media --- p.132 / Chapter 7.2.2 --- Protoplast fusion --- p.133 / Chapter 7.2.3 --- Analyses of progeny --- p.134 / Chapter 7.2.4 --- Phylogenetic analysis --- p.135 / Chapter 7.2.5 --- Fruiting test --- p.135 / Chapter 7.3 --- Results / Chapter 7.3.1 --- Selection of fusion products --- p.135 / Chapter 7.3.2 --- Analyses of fusion progeny --- p.139 / Chapter 7.3.3 --- Identification by protoplasting technique --- p.143 / Chapter 7.3.4 --- Determination of nuclear DNA contents --- p.145 / Chapter 7.3.5 --- rDNA PCR analysis in fusion --- p.148 / Chapter 7.3.6 --- Identification of hybrids by AP-PCR and RAPDs markers --- p.151 / Chapter 7.3.7 --- Phylogenetic analysis --- p.162 / Chapter 7.3.8 --- Fruiting test --- p.164 / Chapter 7.4 --- Discussion --- p.165 / Chapter 7.5 --- Couclusions --- p.169 / Chapter Chapter 8 --- Protoplast fusion in shiitake and other species --- p.171 / Chapter 8.1 --- Introduction --- p.172 / Chapter 8.2 --- Materials and Methods --- p.172 / Chapter 8.3 --- Results and Discussion --- p.173 / Chapter 8.4 --- Couclusion --- p.179 / Chapter Chapter 9. --- General discussion and conclusions --- p.180 / Appendix 1. Determination of ploidy in some mushrooms --- p.187 / Appendix 2. Genomic DNA Isolation --- p.188 / Appendix 3. Arbitrary primer polymerase chain reaction --- p.190 / Appendix 4. rDNA PCR Amplification conditions --- p.193 / Appendix 5. Pulsed Field Gel Electrophoresis --- p.195 / Appendix 6. Genetic distance analysis in hybrids and their parents --- p.196 / References --- p.197

Being the most abundant carbon-containing terrestrial biopolymer, lignocellulose serves as one of the best candidate feedstocks for biofuel production. The current cost-ineffective method for lignocellulose pretreatment is one of the major barriers that hinder the development of biofuel production. This leads to an exploration in the potential application of lignocellulolytic enzymes in this biorefinery process. Taking advantage of the strong activity of ligninolytic enzymes in L. edodes, we aimed at cloning and heterologously expressing these enzymes. The present project applied a yeast expression system, Pichia pastoris, as a laboratory-scale platform for heterologous expression of one of our target ligninolytic enzymes, manganese peroxidase (MnP). We successfully cloned and expressed recombinant MnP. Its enzymatic activity was the highest when grown in the presence of hemoglobin. Our long-term goal is to establish a platform for the large-scale production of recombinant lignocellulotyic enzymes at low-cost, which would strengthen their application in biofuel production. / The shitake mushroom, Lentinula edodes, is one of the most commonly consumed edible mushrooms in Asian countries. It is a saprophyte that naturally colonizes dead wood. As a member of wood-decaying white rot basidiomycete, L. edodes is able to depolymerize lignin and hydrolyze wood polysaccharides. However, the enzymatic mechanism for its lignocellulolytic system is poorly understood. Examination on the L. edodes genome and transcriptome revealed a unique lignocellulolytic system. L. edodes has a diverse enzymatic arsenal for lignin degradation. The enzymes include laccase, manganese peroxidase, cellobiose dehydrogenase and various lignin degrading auxiliary enzymes. When compared to another white rot fungus Phanerochaete chrysosporium, L. edodes possesses more hemicellulase- and pectinase-coding genes, and fewer genes encoding cellulases, suggesting that it preferentially attacks non-cellulosic polysaccharides. The transcription analysis on genes related to antioxidative mechanisms also offers insights to the oxidative stress encountered by mycelium during the free radical-mediated lignin degradation. / Kwok, Sze Wai. / Adviser: Hoi-Shan Kwan. / Source: Dissertation Abstracts International, Volume: 73-01, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2009. / Includes bibliographical references (leaves 141-160). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.