金针菇生活史各阶段核相研究

对金针菇Flammulina velutipes生活史各阶段的菌丝体、子实体、担孢子等进行荧光显微观察。结果表明,金针菇单个担孢子发育而来的同核体菌丝为单核,无锁状联合,部分单孢菌株能形成同核子实体,同核子实体的原担子中只有1个核,该核在担子中进行一次有丝分裂形成纵向排列的2个子核,担子发育停止,同核子实体不产生担孢子;具有可亲和交配型的两个同核体菌丝经过质配形成异核体菌丝,异核体菌丝双核,具有锁状联合,能形成异核子实体,异核子实体的原担子中具有2个核,这2个核经过核配融合为1个二倍核,二倍核在担子中进行减数分裂形成4个单倍子核,4个子核分别进入4个担孢子中,随着担孢子继续发育,其中的单核再进行一次有丝分裂,成熟担孢子为双核,但这2个核是同质的。单核菌丝和双核菌丝都能产生粉孢子,且产生的粉孢子均为单核。     

茯苓基本生物学特性研究

以11个不同来源的茯苓菌株为材料,研究了茯苓菌丝体、子实体和担孢子的形态特征及适宜的生长、发育条件。结果表明,茯苓菌丝体为少分枝、有隔膜、无锁状联合的多核菌丝,茯苓担孢子核相以双核为主,双核孢子,单核孢子和无核孢子分别占87.2%,4.7%和8.1%。配对试验结果表明,同一菌株及不同菌株原生质体分离株间的配对均能融洽生长,同一菌株担孢子间的配对均产生拮抗线,但其中有少数配对在交接区形成扇形区域,拮抗线随后消失,而不同菌株担孢子间的配对则全部形成稳定的栅栏型菌落,暗示茯苓担孢子中的两个细胞核是具遗传互补性,能形成独立个体的异双核,茯苓可能是一种次级同宗结合菌。     

长根小奥德蘑双孢菌株与四孢菌株核相变化的比较

用双苯并咪唑（Hoechst 33258）染色法分别对长根小奥德蘑Oudemansiella radicata双孢菌株和四孢菌株的菌丝、子实体、担孢子进行染色观察,结果表明：双孢长根小奥德蘑菌丝细胞多为单核,无锁状联合;原担子中单核进行一次有丝分裂形成两个横向或纵向排列的子核,这2个子核分别进入2个担孢子中,留下无核的空担子;成熟担孢子具有一个核。四孢长根小奥德蘑菌丝细胞大多数为双核,具有锁状联合;进入原担子中的两个单倍性细胞核先发生核配,形成一个二倍性的核,再经过减数分裂形成四个染色体减半的单倍性子核,     

金针菇粉孢子的生物学研究

采用单孢分离的方法分离培养金针菇粉孢子,对粉孢子的产生、萌发、核相及极性进行了研究。结果表明：单核菌丝和双核菌丝都产生粉孢子,粉孢子多呈圆柱或卵圆形,少数呈圆形或Y形。粉孢子很容易萌发,其芽管直径一般较粉孢子宽。单核菌丝产生单核的粉孢子,其极性与亲本菌丝相同;双核菌丝产生的粉孢子也为单核,未观察到双核或多核的粉孢子。双核菌丝产生的粉孢子一部分与组成双核体的一个亲本单核菌丝的交配型相同,另一部分与组成双核体的另一亲本单核菌丝有相同的交配型。     

黄伞单孢杂交育种的初步研究

以采自山西省五台山和关帝山的WT3 和GD3两个黄伞纯化菌株为材料,显微观察了黄伞担孢子萌发的菌丝及交配特征,在PDA培养基上担孢子首先在其一端萌发出菌丝,宽度约3~4μm,接着产生分枝呈散发状向四周延伸。试验表明:单核菌丝体上可产生类似原基的凸状体,但不具结实性。两个单核菌丝交配后出现锁状联合是形成双核菌丝的典型特征,并具有结实性。     

金针菇粉孢子的生物学研究

采用单孢分离的方法分离培养金针菇粉孢子,对粉孢子的产生、萌发、核相及极性进行了研究。结果表明：单核菌丝和双核菌丝都产生粉孢子,粉孢子多呈圆柱或卵圆形,少数呈圆形或Ｙ形。粉孢子很容易萌发,其芽管直径一般较粉孢子宽。单核菌丝产生单核的粉孢子,其极性与亲本菌丝相同;双核菌丝产生的粉孢子也为单核,未观察到双核或多核的粉孢子。双核菌丝产生的粉孢子一部分与组成双核体的一个亲本单核菌丝的交配型相同。另一部分与组     

利用原生质体融合研究凤尾菇中性菌株的交配反应

凤尾菇营养缺陷型突变型中性菌株与另一株营养缺陷型的正常交配型菌株进行了原生质体融合反应.融合子之间以及与亲本之间在菌落形态和菌丝生长速度上表现出了较大差异。融合子菌落角变的菌丝体从多核体转变为双核体,并且在菌丝体上产生了圆桶状的特殊结构而不是通常的锁状联合。初步观察表明这些结构似乎与细胞核的迁移有关.对融合子子实体的担孢子进行了遗传分析,结果确证子实体是由两个亲本菌株的融合子产生,同时也表明要对中性菌株交配反应中基因重组进行深入分析还需要进行更多的研究。     

利用原生质体融合研究凤尾菇中性菌株的交配反应

凤尾菇营养缺陷型突变型中性菌株与另一株营养缺陷型的正常交配型菌株进行了原生质体融合反应融合子之间以及与亲本之间在菌落形态和菌丝生长速度上表现出了较大差异。融合子菌落角变的菌丝体多核体转变为双核体,并且在菌丝体上产生了圆桶状的特殊结构而不是通常的锁状联合。初步观察表明这些结构似乎与细胞核的迁移有关,对融合子子实体的担孢子进行了遗传分析,结果确实了实体是由两个亲本菌株的融合子产生,同时也表明要对中性菌株交配反应中基因重组进入深入分析还需要进行更多的研究。     

木耳和毛木耳的极性研究

从木耳(Auricularia auricula)和毛木耳(A.polytricha)的同一子实体弹射、分离30个单孢子并发育成单核菌丝体,各自分成3组,以10×10方式进行单核体两两配对。取两配对单核体交结处菌丝体块到新的平板上继续发育并插入无菌的盖玻片让其菌丝爬上。后利用双苯并咪唑(Hoechst 33258)染色,在萤光显微镜下逐块检查配对后菌丝体细胞中核的数目。如果出现双核,再加以检查锁状联合以验证,则为配对亲和。不亲和者仍为单核。根据配对行为进行不亲和因子分配决定其交配型。检测结果表明,木耳和毛木耳担孢子的性别是由一对遗传因子A.a所控制。属典型性二极性(bipolar)异宗结合。     

木耳和毛木耳的极性研究*

从木耳(Auricularia auricula)和毛木耳(A.polytricha)的同一子实体弹射、分离30个单孢子并发育成单核菌丝体,各自分成3组,以10×10方式进行单核体两两配对。取两配对单核体交结处菌丝体块到新的平板上继续发育并插入无菌的盖玻片让其菌丝爬上。后利用双苯并咪唑(Hoechst 33258)染色,在萤光显微镜下逐块检查配对后菌丝体细胞中核的数目。如果出现双核,再加以检查锁状联合以验证,则为配对亲和。不亲和者仍为单核。根据配对行为进行不亲和因子分配决定其交配型。检测结果表明,木耳和毛木耳担孢子的性别是由一对遗传因子A.a所控制。属典型性二极性(bipolar)异宗结合。     

Mating System and Basidiospore Formation in the Lignin-Degrading Basidiomycete Phanerochaete chrysosporium

Prototrophic strains recovered from crosses between auxotrophic strains of the lignin-degrading basidiomycete Phanerochaete chrysosporium were induced to fruit. The progeny of most of these self-crosses were prototrophic, indicating that the nuclei of the original prototroph were wild-type recombinants rather than complementary heterokaryons and that the binucleate basidiospores of this organism are homokaryotic. Various wild-type strains were shown to have multinucleate cells lacking clamp connections and to possess a variable number of sterigmata per basidium. Colonies arising from single conidia of various wild-type strains were all capable of producing fruit bodies and basidiospores. In addition, single basidiospores from three wild-type strains all produced fruit bodies and basidiospores. Nonfruiting as well as fruiting isolates were obtained from single basidiospores of five other wild-type strains. Basidiospores from these fruiting isolates always yielded colonies that fruited, again indicating that the spores are homokaryotic. Nonfruiting isolates from the same strain did not produce basidiospores when allowed to form a heterokaryon, implying that these isolates do not represent mating types. All this evidence indicates that P. chrysosporium has a primary homothallic mating system. In addition to fruiting and nonfruiting phenotypes, basidiospores from strain OGC101, a derivative of ME-446, gave rise to colonies which did not grow on cellulose (Cel⁻). The fruiting, nonfruiting, and Cel⁻ phenotypes differed from each other and from the parental wild-type strain in a variety of characteristics, including growth, conidiation, and evolution of ¹⁴CO₂ from ¹⁴C-side chain-labeled lignin, indicating that strain OCG101 is a heterokaryon.     

Establishment of monokaryotic and dikaryotic isolates of Hedgehog mushrooms (Hydnum repandum and related species) from basidiospores

Hydnum repandum and its relatives are gourmet edible ectomycorrhizal mushrooms. However, no reliable pure cultures have been reported in this genus. Here, we report for the first time the successful isolation of mycelial strains from basidiospores in the genus Hydnum. Basidiospores obtained from basidioma samples were aseptically inoculated onto modified Norkran's C (MNC) medium, MNC containing n-butyric acid (n-MNC), or MNC with gellan gum instead of agar (G-MNC). Although basidiospore germination was observed in most samples, the isolation rate was higher from MNC (91.7%) and G-MNC (93.8%) than from n-MNC (36.4%). Most established isolates were monokaryotic and lacked a clamp connection, but three were dikaryotic and had clamp connections. Established isolates were identified by molecular phylogenetic analysis of the internal transcribed spacer region of the rRNA gene. These results suggest that basidiospores can be used to establish monokaryotic and dikaryotic isolates of Hydnum species.     

桦纤孔菌有性生殖相关特征及交配型位点分析

对桦纤孔菌菌株MDJCBS88的显微形态、菌丝及担孢子核相进行了观察。采用棉籽壳培养基对担孢子萌发形成的菌株进行栽培试验,筛选出不形成子实体或子实体发育不完整的菌株,将这些菌株在平板上进行了亲和试验,分析桦纤孔菌的有性生殖方式;并基于基因组序列进行交配型基因克隆验证,分析桦纤孔菌的交配型位点结构。显微观察发现,桦纤孔菌菌丝没有锁状联合结构,菌丝细胞无核到多核;子实层担孢子可含0-4个不等的细胞核,不同时期弹射的担孢子含有的细胞核数量不同。桦纤孔菌担孢子萌发率极低,能萌发的担孢子多为早期弹射的担孢子;培养基也影响担孢子的萌发率,与PDA培养基和CYM培养基相比,桦木屑培养基最适合桦纤孔菌担孢子萌发,萌发率为4.55%。从担孢子萌发的96个菌株中获得了2个不结实菌株和9个结实不产孢菌株,占11.5%,这些菌株间亲和试验出现不同的表现特征,包括形成产孢子实体,产生菌丝纽结,相互融合和相互拮抗等现象,认为桦纤孔菌的有性生殖以次级同宗结合为主,并受交配型基因控制。交配型位点克隆测序后分析发现,桦纤孔菌交配型A位点共14 034 bp,含有一个MIP基因和两组HD1和HD2基因;交配型B位点包含3个疑似信息素受体基因和1个信息素前体编码基因。     

Alternation of nuclear phase in the filamentous basidiomycete,Helicobasidium mompa

Variation in the number of nuclei and cellular ploidy were observed in eight strains ofHelicobasidium mompa. The basidiospores, single-spore isolates and field-isolated strains were all dikaryons. The cellular ploidy, which was assessed by analyzing the fluorescence emitted by DAPI-stained nuclei, was unstable: monokaryotic strains derived from the original dikaryotic strains by successive subcultures were mainly tetraploid, although the original dikaryon was in most cases diploid. On the other hand, a dikaryotic strain derived by treatment with benomyl was haploid. These results suggest that diploid dikaryon is a normal nuclear phase ofH. mompa in nature, and the alternation of ploidy may be due to a feature of the mating system of this fungus.     

The cell type ofRhodosporidium toruloides after protoplast fusion between strains of identical and opposite mating type

Protoplast fusions between strains of identical and of opposite mating type were performed. Sexual crosses and protoplast fusions inRhodosporidium toruloides led to different hybrid types. Sexual crosses gave rise exclusively to a dikaryotic mycelium. In protoplast fusions between strains of identical mating type (A ora), monokaryotic yeast-like hybrids which sustained the parental mating type were obtained. In protoplast fusions between strains of opposite mating type, the majority of the hybrids belonged to theMyc ⁺ type, i.e., the hybrids grew like yeasts, able to switch over spontaneously to mycelial growth. In addition to theMyc ⁺ types, up to 10% real yeast hybrids and less than 5% dikaryotic mycelia were obtained. Obviously the cell type inR. toruloides is under the control of the mating type alleles.     

Isolation of a homothallic mutant in <Emphasis Type="Italic">Lentinula edodes</Emphasis>

To obtain a homothallic mutant in Lentinula edodes, basidiospores derived from the common Bmut dikaryon (A1B1mut × A2B1mut) were treated with UV irradiation. Of a total of approximately 5000 monosporous cultures recovered, a single basidiospore isolate was found to produce the hyphae bearing clamp connections without mating. This mutant strain could form fruit bodies, and all its single basidiospore isolates developed into colonies with clamp connections. Such homothallic behaviors were transmitted from the mutant strain to the next generation. During the germination and following hyphal elongation in a single basidiospore of mutant strain, clamp connections were clearly detected in multicellular hyphae, which contained two nuclei in each cell. Their clamp connections were morphologically variable, viz., pseudo, abnormal, and true clamps. Amplified fragment length polymorphism (AFLP) profiles among the basidiospore isolates of mutant strain were identical, indicating that the mutant strain produced isogenic basidiospore progeny. Contribution no. 385 from the Tottori Mycological Institute