Internuclear Genetic Transfer in Vegetative Dikaryons of SCHIZOPHYLLUM COMMUNE: I. Di-Mon Mating Analysis

A series of hemi-compatible dikaryon x monokaryon (di-mon) matings was designed to determine whether there was any genetic interaction between the dikaryotic nuclei. One of the nuclei in each dikaryon was known to carry a recessive gene (mnd) that promoted the development of an abnormal growth form, mound. Dikaryons containing both mnd ⁺ and mnd nuclei produced mosaic colonies that consisted of three distinct kinds of hyphae: mound, fruiting body, and vegetative (devoid of any multihyphal structure). When dikaryotic hyphae from each of these morphological regions were used in di-mon matings, the genetic and developmental characteristics of the selected nuclear types were examined in the resulting derived dikaryons. The results showed that fruiting-body and vegetative cells contained the expected mnd and mnd⁺ nuclei. Dikaryotic mound hyphae, however, contained only mnd nuclei. In a manner as yet unresolved, but clearly dependent on the presence of the mnd allele, the mnd ⁺ allele of a wild nucleus was altered to, or acquired, the mnd allele. A number of hypotheses were considered to explain the genetic event(s) that generates [mnd + mnd*] dikaryotic cells from [mnd⁺ + mnd] cells, but none was found to be entirely satisfactory.     

Sexual Incompatibility Factors and Somatic Recombination in SCHIZOPHYLLUM COMMUNE

Two sets of diploid cultures of S. commune were observed for sectoring due to haploidization or recombination. Each set consisted of compatible and common-AB diploids otherwise almost isogenic. One of the sets included two compatible diploids with a large proportion of dikaryotic cells. The sectors were isolated and analysed for evidence of aneuploidy and frequent crossing over to determine whether they arose via mitotic or meiotic-like events. It was found that the recombination process in both common-AB and compatible diploids was predominantly mitotic. However, the compatible diploids which developed a high frequency of dikaryotic components gave some evidence of meiotic-like activity. Thus, compatible mating-type factors are necessary for dikaryosis, but not sufficient in themselves to produce it. In compatible mycelia where dikaryosis does occur, meiotic-like recombination may also occur. It is proposed that both lapse into the dikaryotic state, and meiotic-like recombination was induced by different genes under control of the incompatibility factors. Dikaryosis and meiosis are thus seen as tandem phenomena, neither causal of the other but both induced by action of compatible mating-type factors.     

Genetic Control of Recombination in SCHIZOPHYLLUM COMMUNE: Location of a Gene Controlling B-Factor Recombination

In S. commune the frequency of recombination between the two subunits, α and β, of the B incompatibility factor is genetically controlled. Analysis of the progeny obtained from crosses between high- and low-recombining strains indicates that the gene controlling recombination frequency in the B factor is linked to the B factor itself, approximately nine map units from Bβ. This gene, called B-rec-1, does not affect the recombination frequency in an unlinked region (between the subunits of the A incompatibility factor) or in a region contiguous with the B factor (between Bα and the morphological marker dome-2).     

POLYOLS IN SCHIZOPHYLLUM COMMUNE

Sugars and sugar alcohols present in extracts of the wood-rotting mushroom Schizophyllum commune were identified by paper chromatography during fruiting, basidiospore germination, and growth of vegetative mycelium. Homokaryotic fruitbodies and dikaryotic fruits derived from several compatible matings of S. commune contained mannitol and arabitol. Basidiospores shed from dikaryotic fruits also contained mannitol and arabitol while the latter disappeared during spore germination. Vegetative mycelium (strain 699) contained glucose, fructose, mannitol and glycerol while these compounds as well as arabitol occurred in mycelium of strain 845. Polyols are not, therefore, associated exclusively with the sporulation process in S. commune.     

Internuclear Genetic Transfer in Dikaryons of SCHIZOPHYLLUM COMMUNE. II. Direct Recovery and Analysis of Recombinant Nuclei

The recessive gene, mound (mnd), allows the appearance of globose masses of compacted hyphae. Dikaryons of Schizophyllum commune that are heteroallelic for mnd [(mosaic dikaryons: (mnd + mnd(+))] have been successfully dedikaryotized in cholate-containing medium in order to recover the component nuclear types directly. The relative proportion of the two recovered monokaryotic types shows in all cases a marked deviation from 1:1. Hyphae from nonmound mycelial regions yield monokaryotic types identical to those originally used to form the dikaryons. In hyphae from mound-forming regions, however, homoallelism of the mnd allele has been demonstrated; the nuclear type that formerly contained the mnd(+) allele acquired a mnd allele.-The process of internuclear transfer or recombination is unaccompanied by the simultaneous alteration of any additional genetic markers carried by the recipient nucleus. The newly acquired mnd allele segregates in Mendelian fashion in subsequent outcrosses and appears to be chromosomally located. A novel process of somatic recombination, with several features distinct from classical parasexual mitotic recombination, appears to be in operation.     

MUTATIONS AFFECTING HETEROKARYOSIS IN SCHIZOPHYLLUM COMMUNE

Mutations that affect the basic characteristics of heterokaryons of S. commune occur spontaneously and are preferentially selected in the common-A heterokaryon and in its homokaryotic mimics, strains carrying a mutated B factor or strains disomic for heteroallelic B factors. Nine independent mutations were compared: all segregate independently of A and B incompatibility factors, and at least 3 distinct loci, of which 2 are linked, are involved. None of the mutations is phenotypically expressed in the homokaryon or in the common-AB heterokaryon. All 9 mutations increase vegetative vigor of the common-A the effects of all the mutations are additive in both heteroallelic and homoallelic combinations. At least 1 type-II mutation also affects nuclear distribution in common-B heterokaryons. Type-II mutations appear to reduce common-A, common-B, and compatible heterokaryons to a single type unlike any of the normal heterokaryons. Pseudoclamping often persists for extended periods in modified homokaryons isolated from modified heterokaryons. Several cases of somatic recombination have been observed among components of modified heterokaryons.     

EVIDENCES OF COMMON-AB HETEROKARYOSIS IN SCHIZOPHYLLUM COMMUNE

Since preliminary experimental evidence for the existence of common-AB heterokaryons in Schizophyllum commune was inconclusive, experiments were performed to test for and to characterize these heterokaryons. Prototrophic mycelia regularly result from the growing together of pairs of mutant strains of identical mating type which carry non-allelic nutritional deficiencies. Since crossfeeding through a dialyzing Cellophane membrane does not occur, the prototrophic common-AB mycelia apparently have both parental nuclear types within a common cytoplasm. Hyphal tips isolated from the peripheries of these prototrophic mycelia usually are not prototrophic. The distributions of the parental nuclear types in the subcultured prototrophic mycelia, as revealed by grid sampling, are irregular in pattern and extent; these patterns probably reflect the nuclear ratios in the transfer inocula, as well as the distributions of nuclear types in various parts of the inocula. Since a common-AB prototroph typically consists of regions containing a single nuclear type as well as regions containing both nuclear types, marked fluctuations of the estimated nuclear ratio occur upon subculture, and many small transfer-inocula are not prototrophic as they contain only a single auxotrophic nuclear type. The patterns of nuclear distributions in prototrophic common-AB mycelia are probably maintained by the restriction of nuclear migration.     

NUCLEAR MIGRATION IN DIKARYOTIC-HOMOKARYOTIC MATINGS IN SCHIZOPHYLLUM COMMUNE

The present data indicate internuclear selection in dikaryotic-homokaryotic matings is not due to differential rates of migration of the 2 types of migrant nuclei or to differential rates of saturation of the homokaryon. The advancing fronts of the 2 types of migrating nuclei are separated by essentially the same distance shortly after migration has been initiated and during later periods in the migration process. Internuclear selection, controlled by the incompatibility factors, must occur during the initiation of migration or earlier. Migration of both types of nuclei from the dikaryon has been demonstrated in hemicompatible and incompatible matings.     

A Class of Genes Affecting B Factor-Regulated Development in SCHIZOPHYLLUM COMMUNE

Twelve mutations affecting nuclear migration, a major developmental phase in Schizophyllum commune, display a complex pattern of complementation and recombination. They are expressed only when a genetic factor controlling this phase of development, the B incompatibility factor, is operative. All twelve mutations are linked to the B factor, nine in a cluster and three in distinct loci outside the cluster. A linear map cannot be constructed from the frequency of recombination. Complementation maps are not linear. There is little correlation between the complementation groups and the groups based on recombination. Many pairs of mutations that do not complement recombine with frequencies of 1.1% to 26.9%. The genes represented by the twelve mutations are located in a linked group of about 18 known genes involved in the specific phase of development controlled by the B factor.     

凤尾菇和裂褶菌原生质体非对称融合研究

具有双重营养缺陷型的裂褶菌单核体突变菌株的原生质体经过灭活处理后作为供体与同样带有营养缺陷型标记的风尾菇单核体原生质体融合,得到大量生长速度和菌落形态差异较大的多核体和双核体融合子．这些融合子主要显示了供体菌株的特性,数次转接后,有的融合子从多核体转变成双核体,有的融合子完全恢复了供体亲本双核体的遗传特性,同时获得了一些有意义值得继续探讨的新的生理特性,如菌丝体在木屑 皮以及废棉培养料上生长速度较快,在平板培养基和木屑培养料上均较早较快地形成原基和子实体等．结果表明原生质体非对称融合是一种效率较高的融合方式,可以作为食用菌育种的一种有效途经．     

Complementation and Noncomplementation among Nonallelic Mutations Altering Development in SCHIZOPHYLLUM COMMUNE

Mutations resistant to compounds that alter the development of Schizophyllum commune (cyclic adenosine monophosphate, caffeine and imidazole) were selected. These mutations defined four genes, car (cAMP resistance), caf-1 and caf-2 (caffeine resistance) and imr (imidazole resistance). The mutations were found to be noncomplementing in certain pairs when tested against each other and against two previously isolated developmental mutations, B-con and sep (septateless), in fully compatible, forced heterokaryons. The pattern of complementation in fully compatible diploids showed noncomplementation only between allelic mutations, except for one case, imr vs. car.     

New Mutations and a 7-Chromosome Linkage Map of SCHIZOPHYLLUM COMMUNE

Forty-eight useful new mutations of S. commune were obtained by mutagenesis with N-methyl-N'-nitro-N-nitrosoguanidine. Their requirements and meiotic linkage relationships to each other and previously mapped areas were investigated. Several of these new mutations were incorporated into diploid strains so that the diploids contained at least one marker on every linkage group. Analysis of somatic recombination in these diploids indicated that each meiotic linkage group corresponded to an independent chromosome.     

裂褶菌深层培养及多糖测定

为了深层培养裂褶菌Schizophyllum commune Fr.产生多糖,对产多糖的适宜培养基,最佳时间,高产菌株进行了研究。从南京灵谷寺及南京大学校园生长的裂褶菌子实体分离到3株产多糖的裂褶菌菌株,编号南大835,南大843,南大853。对南大843用6种不同培养基进行深层培养,测定和比较了多糖和菌丝产量,其结果表明黄豆粉葡萄糖液体培养基是适于裂褶菌合成多糖的培养基,能培养出密集、白色、均匀的菌球和丰富的多糖。其组成为(g/L):葡萄糖30,黄豆粉5,酵母膏2,KH_2PO_4 1,MgSO_4·7H_2O0.5。pH5.5。最适发酵条件:pH5—5.5,温度26—28℃,振速:100—110次/分,当pH降至4.9—4.7,残糖量在1%以下,5—6天可终止发酵。在培养6天的浓缩滤液中加入等体积的95%乙醇后大量白色粘稠、纤维状的多糖被沉淀下来。在上述发酵条件下,3个菌株比较结果,南大853能明显提高多糖产量,6天的培养液中多糖量可达5.5—6g/L,南大843和南大835分别是5g/L和2.8g/L。     

深层培养裂褶菌胞外多糖的提取及结构研究

对深层培养裂褶菌 (Schizophyllumcommune)胞外多糖的提取工艺及多糖结构进行了初步研究。将等电点法与Sevag法相结合可高效的去除多糖中的蛋白 ,其方法简单有效。纯多糖经凝胶柱层析 ,聚丙烯酰胺凝胶电泳 ,高效液相色谱分析为均一组分 ,分子量 4×1 0 4D。通过完全水解 ,纸层析 ,气相色谱分析单糖组分 ,红外光谱 ,酶解反应 ,高碘酸氧化分析结构 ,证明了裂褶菌多糖是以葡萄糖为单一组分 ,β (1 3)和β (1 6)糖苷键组成的β D葡聚糖。