Hybridisation,genomic constitution and generic delimitation inElymus s. l. (Poaceae: Triticeae)

Intergeneric crosses were made between representatives of the genomically-defined generaElymus, Agropyron, Elytrigia, Pseudoroegneria, andThinopyrum. The genomic constitution ofElytrigia repens, the type species ofElytrigia, is shown to be SSH, a genomic combination otherwise found only inElymus. The S genome ofPseudoroegneria has almost always a dominant influence on the morphology of the taxa of which it is a component.Wang (1989) showed that the J genome inThinopyrum and the S genome have considerable homoeology, with a mean c-value of 0.35 in diploid SJ hybrids. A genetic coherence from S to SJ^e, J^e, J^eJ^b, and J^b can be expected, agreeing with the continuous morphologic variation pattern observed. Because of the absence of morphological discontinuities between the taxa,Pseudoroegneria (S),Elymus (SH, SY, sometimes with additional genomes),Elytrigia (SSH, SSHX), andThinopyrum (SJ, SJJ, J) are best treated as a single genus,Elymus, following the generic concept ofMelderis in Flora Europaea and Flora of Turkey. The basic genomic constituents ofElymus will then be the S and/or J genomes.Agropyron, with diploids, tetraploids, and hexaploids based on the P genome is morphologically distinct from other genera inTriticeae. In a few species ofElymus andPseudoroegneria, a P genome is an additional constituent. In these cases the P genome has a negligible morphological influence. Therefore, it seems reasonable to maintainAgropyron as a separate genus.     

Genome analysis of species in the genus Hystrix (Triticeae; Poaceae)

Genomic in situ hybridisation (GISH) and Southern genomic hybridisation were applied in order to gain further knowledge regarding generic delimitation of the genus Hystrix as well as to clarify the genomes of the Hystrix species H. patula, H. longearistata, H. coreana, H. duthiei and H. komarovii. The hybridisation intensity of different genomic probes was compared among the Hystrix species and with other Triticeae species. The Southern- and GISH results confirm that H. patula contains the StH genome and show that H. komarovii most likely has a variant of this StH genome. The other Hystrix species under study, i.e. H. longearistata, H. coreana and H. duthiei, contain an Ns basic genome, and most probably two variants of this basic genome, Ns ¹ Ns ² . The genus Hystrix is thus not a monophyletic group of species.     

垂穗披碱草与达乌里披碱草基因组细胞遗传学比较

采用顺序FISH-GISH技术,12个重复序列探针,包括9个三核苷酸简单重复序列、2个卫星DNA重复序列pSc119.2和pAs1以及5S rDNA,通过重复序列的物理定位对达乌里披碱草和垂穗披碱草基因组中部分重复序列的分布特征进行了比较分析,为进一步研究垂穗披碱草和达乌里披碱草的物种形成及演化提供新的分子细胞遗传学证据。结果表明:(1)所有的序列在这2个物种的染色体上都能产生可检测的杂交信号,且在2个物种中(AAC)_(10)、(ACT)_(10)、(CAT)_(10)都表现为共分布,(AAG)_(10)与(AGG)_(10)表现为近似共分布;2个物种的H基因组除5S rDNA序列外,其他序列都产生强烈且丰富的杂交位点,St与Y基因组不同重复序列探针的荧光位点数目有所差别,表现为5S rDNA、pSc119.2、(AAC)_(10)、(CAT)_(10)、(ACT)_(10)、(CAC)_(10)探针的信号位点较少或无信号,其余的探针信号位点稍多。(2)达乌里披碱草的第2对染色体上具有(AAC)_(10)、(CAT)_(10)、(ACT)_(10)的杂交位点、第6对染色体上具有(CAC)_(10)的杂交位点,而在垂穗披碱草的St基因组中未观察到上述序列杂交位点;达乌里披碱草St基因组仅有第4对染色体的端部具有pSc119.2杂交位点,而在垂穗披碱草St基因组中的pSc119.2杂交位点位于第5对染色体长臂的间隔区;相对于达乌里披碱草,垂穗披碱草St和Y基因组染色体含有更多的重复序列杂交位点。(3)达乌里披碱草的H/Y基因组间易位在不同材料间是稳定存在的,达乌里披碱草基因组相对稳定,不同材料间H基因组重复序列杂交信号多态性高于St和Y基因组;垂穗披碱草基因组的变异较大,不同材料间St和Y基因组重复序列杂交信号多态性高于H基因组。研究认为,垂穗披碱草和达乌里披碱草的H基因组均起源于布顿大麦,St基因组可能起源于不同的拟鹅观草属物种;与达乌里披碱草相比垂穗披碱草St与Y基因组可能具有更高的染色体结构变异性,而垂穗披碱草St与Y基因组变异较大的原因可能是与同区域分布的含StY基因组的物种发生了种间渗透杂交。     

Asymmetrical development of biovulate cones resulting in uniovulate cones in Ephedra rhytidosperma (Ephedraceae)

Female cone morphology in Ephedra, including the number of initiated ovules and mature seeds per cone, provides important taxonomic characters used in sectional or species delimitation within Ephedra. Recent molecular phylogenies have indicated, however, that seed number per cone has changed repeatedly during the evolution of the genus. This study reports on the development of the female cone of E. rhytidosperma, based on a large sample of dissected cones studied under SEM. All cones were initially biovulate, and in the majority of cases, both female reproductive units (FRUs) developed a micropylar tube and formed mature seeds. In a few cases, the FRU pair developed asymmetrically in a cone, with one of them eventually aborting. There was no evidence of fusion of the FRU pair. Phylogenetically, E. rhytidosperma is in a clade with E. equisetina, which has uniovulate cones, and E. gerardiana and E. minuta, which have biovulate cones that also become unispermous via abortion. The biovulate condition may thus be ancestral in this clade.     

Electrophoretic karyotypes of some related Mucor species

Contour clamped homogeneous electric field (CHEF) gel electrophoresis was used to obtain electrophoretic karyotypes from nine Mucorstrains representing five different species (M. bainieri, M. circinelloides, M. mucedo, M. plumbeus and M. racemosus). The chromosomal banding patterns revealed high variability among the isolates. The sizes of the DNA in the Mucor chromosomes were estimated to be between 2.5 and 8.7 Mb. The total genome sizes were calculated to be between 30.0 and 44.7 Mb. The applicability of these electrophoretic karyotypes for the investigation of genome structure, for strain identification and for species delimitation is considered.     

Relationships betweenOryza species (Poaceae) based on 5S DNA sequences

Relationships between 9Oryza species, covering 6 different genomes, have been studied using hybridization and nucleotide sequence information from the5S Dna locus. Four to five units of the major size class of 5S DNA in each species, 55 units in all, were cloned and sequenced. Both hybridization and sequence data confirmed the basic differences between the A and B, C, D genome species suggested by morphological and cytological data. The 5S DNA units of the A genome species were very similar, as were the ones from the B, C, and D genome-containing species. The 5S DNA ofO. australiensis (E genome) grouped with the B, C, D cluster, while the units ofO. brachyantha (F genome) were quite different and grouped away from all other species. 5S DNA units fromO. minuta, O. latifolia, O. australiensis, andO. brachyantha hybridized strongly, and preferentially, to the genomic DNA from which the units were isolated and hence could be useful as species/genome specific probes. The 5S DNA units fromO. sativa, O. nivara, andO. rufipogon provided A genome-specific probes as they hybridized preferentially to A genome DNA. The units fromO. punctata andO. officinalis displayed weaker preferential hybridization toO. punctata DNA, possibly reflecting their shared genome (C genome).     

Analysis of metaphase I chromosome association in species of the genus Aegilops

Summary Metaphase-I chromosome associations in every diploid and polyploid species of the genus Aegilops were studied using C-banding in order to analyse the cytogenetic behaviour of the whole complement as well as of specific genomes in different polyploid species. Differences were observed in the frequency of associations per cell among different species of the same ploidic level and even between species sharing the same genomic constitution. Differences were also found between different genomes within the same polyploid species and between the same genome when present in several diploid and polyploid species. Several factors proposed as having an influence on the frequency of metaphase-I associations, such as chromosome morphology, C-heterochromatin content, genetic control and genome interactions, are discussed. Most of the polyploid Aegilops species showed a diploid-like behaviour at metaphase I although multivalents involving homoeologous associations were occasionally observed in Ae. biuncialis, Ae. juvenalis and Ae. crassa(6x); therefore, the Aegilops diploidising genetic system is not equally effective in all polyploid species.     

Enhancement of sporulation in species of Bipolaris, Curvularia, Drechslera, and Exserohilum by growth on cellulose-containing substrates

Nine species of Bipolaris, Curvularia, Drechslera, and Exserohilum were compared for sporulation on agar media and for enhancement of sporulation by growth on four cellulose-containing substrates (index card, filter paper, cheesecloth, cotton fabric). On two natural and one synthetic agar media, sporulation varied from profuse to nonexistent among three isolates of each species. Growth of all species on cellulose substrates resulted in large and significant increases in sporulation. Growth on index card pieces often provided the greatest increases, but no single substrate was superior for all species, and significant substrate × isolate interactions were observed within species. Overlay of filter paper onto whole colonies in agar plates resulted in 2 to 18-fold increases in sporulation for eight of nine species and production of spores in sufficient quantity for most experimental purposes. Overlay of soil dilution plates with filter paper to promote sporulation of colonies enabled detection of B. spicifera, B. hawaiiensis, C. lunata, and E. rostratum at relatively low population levels (≤1.3 × 10³ colony-forming units per gram of soil) in samples of a naturally infested soil. Results indicate that enhancement of sporulation by growth of species of Bipolaris, Curvularia, Drechslera, and Exserohilum on cellulose substrates may facilitate (i) their identification in culture, (ii) production of spores at relatively high concentrations, and (iii) detection and enumeration of these fungi in soil.     

Evidence of widespread hybridization among couch grasses (Elymus,Poaceae)

Hybridization, polyploidization, and crop‐to‐wild gene transfer within the agriculturally important tribe Triticeae are well explored experimentally, but the true consequences of both phenomena under natural conditions remain understudied. The present paper reports on an investigation of three species of couch grasses (Elymus hispidus, E. repens, and E. caninus) examining the ploidy levels and absolute genome sizes (1081 plants from 302 natural populations in Central Europe, verified by chromosome counts) and their morphological delimitation. In the present study, the hexaploid level prevailed in E. hispidus and E. repens whereas E. caninus was exclusively tetraploid. Introgressive hybridization between hexaploid species, unidirectionally shifted towards E. hispidus, was indicated by a continual pattern of genome size values. We did not find any evidence for heteroploid hybridization involving tetraploid E. caninus; however, we detected minority cytotypes among both E. caninus plants (hexaploid) and E. repens–E. hispidus hybrids (heptaploid and nonaploid) suggesting the formation of unreduced gametes. Morphometric results (367 plants, redundancy analysis, principal component analysis, and correlation analysis) mirrored the continual homoploid pattern of absolute genome size (including the unidirectional shift), and a significant correlation between absolute genome size and morphology was confirmed. Moreover, morphometric analyses detected additional characteristics for the delimitation of the Elymus taxa under study. Considering the crossability of E. hispidus with Triticum aestivum (bread wheat), the revealed extent of introgressive hybridization has implications for assessing the potential risk of gene flow between crops and troublesome weeds.     

Molecular diversity of species of the Elymus trachycaulus species complex and their relationships to non-North American taxa

Species of the E. trachycaulus complex species are known for their morphological variability, but little is known about their genetic basis. The delimitation of taxa within the complex has been controversial and difficult. E. trachycaulus is predominantly self-pollinating, and lacks clear morphological boundaries between it and E. alaskanus. Another controversial taxonomic issue of E. trachycaulus is the relationships of this complex species to non-North American E. caninus. The objectives of this study were to examine genetic diversity and the systematic relationships among the species of the E. trachycaulus complex and their relationships with E. caninus, E. alaskanus and E. mutabilis. Random amplified polymorphic DNA method was used to study 35 accessions of E. trachycaulus complex and other Elymus species. Higher genetic variation was detected within species of E. trachycaulus complex. Eurasian accessions are as variable as the North American ones. Both UPGMA and NJ analyses did not show clearly separation among species of the E. trachycaulus complex. No clear association between geographic origin and genetic grouping among these species was found. Eurasian E. trachycaulus probably originated from multiple North American populations.     

Identification of genome constitution of Oryza malampuzhaensis, O. minuta,and O. punctata by multicolor genomic in situ hybridization

Multicolor genomic in situ hybridization (McGISH) was applied to identify the genomic constitution of three tetraploid species (2n = 4x = 48) in the Oryza officinalis complex of the genus Oryza, i.e. Oryza malam-puzhaensis, Oryza minuta, and Oryza punctata. The genomic probes used were from three diploids, i.e. Oryza officinalis (CC), Oryza eichingeri (CC) and Oryza punctata (BB), respectively. The results indicated that all three tetraploids are allotetraploid with the genomic constitution of BBCC, and among them the genome constitution of O. malampuzhaensis was verified for the first time. Restoration of the independent taxonomic status of O. malampuzhaensis is suggested. One pair of satellite chromosomes belonging to the B genome was identified in O. malampuzhaensis, but no such satellite chromosomes were found in either O. minuta or the tetraploid O. punctata. The average chromosome length of the C genome was found to be slightly larger than that of the B-genome chromosomes of O. minuta, but not in the tetraploids O. punctata and O. malampuzhaensis. McGISH also revealed that the B genome of O. minuta and the B genome of diploid O. punctata showed clear differentiation from each other. Therefore, the suggestion was proposed that the B genome in diploid O. punctata was not the source of the B genome of O. minuta. The present results proved that multicolor GISH had high resolution in identifying the genomic constitution of polyploid Oryza species. Received: 14 February 2000 / Accepted: 13 November 2000     

Relationships between species ofLeymus,Psathyrostachys, andHordeum (Poaceae,Triticeae) inferred from Southern hybridization of genomic and cloned DNA probes

We have used total genomic DNA as a probe to size-fractionated restriction enzyme digests of genomic DNA from a range ofTriticeae species from the generaLeymus Hochst.,Psathyrostachys Nevski, andHordeum L., and hybrids betweenHordeum andLeymus to investigate their taxonomic relationships. Genomic Southern hybridization was found to be an effective and simple way to assess the distribution and diversity of essentially species-specific and common, repetitive DNA sequences, and is hence especially useful in evolutionary studies. The DNA sequences ofH. vulgare seem to diverge substantially from those ofH. brachyantherum, H. lechleri, H. procerum, andH. depressum. The genome ofThinopyron bessarabicum shows little homology to those of theLeymus species investigated, confirming thatT. bessarabicum is not an ancestral genome inLeymus. Although the genomes ofLeymus andPsathyrostachys share substantial proportions of DNA sequences, they include divergent repeated sequences as well. Hybridization with a ribosomal DNA probe (pTa 71) showed that the coding regions containing structural genes encoding the 18 S, 5.8 S, and 26 S ribosomal RNA were conserved among the species investigated, whereas the intergenic spacer region was more variable, presenting different sizes of restriction fragments and enabling a classification of the species. The rye heterochromatin probe pSc 119.2 hybridized to DNA fromH. lechleri andT. bessarabicum, but not to DNA from the other species investigated.     

Genetic variability and genomic divergence of Elymus repens and related species

Summary In order to study the genetic variation and phylogenetic relationship in Elymus repens, amplified fragment length polymorphism (AFLP) were used, together with sequence data for the nuclear gene phytochrome B, phyB, and the chloroplast ribosomal protein encoding gene rps4. A total of 83 collected E. repens samples, 3 E. repens reference samples and 18 related species accessions were analysed and compared with 13 GenBank sequences. AMOVA analysis revealed a moderate genetic differentiation between the populations of E. repens in the three Swedish provinces investigated, while no differentiation was observed due to landscape type. A moderate genetic differentiation was also found when samples from different fields in one province were compared to samples from a selected field. A common female origin was found in E. repens and seven other Elymus species, Pseudoroegneria spicata, Thinopyrum intermedium, T. junceum, Hordeum bogdanii and H. stenostachys. The latter two both harbour the H genome. Taken together, the data suggest that the Swedish E. repens population is slightly heterogeneous and comprises multiple origins of genome donors; a nuclear genome with contributions from Pseudoroegneria (St), Hordeum (H), Thinopyrum (E) and Y with an unknown donor together with a maternal genome donated from Pseudoroegneria.     

Phylogenetic analysis of questionable tetraploid species in Roegneria and Pseudoroegneria (Poaceae: Triticeae) inferred from a gene encoding plastid acety1-CoA carboxylase

To evaluate the phylogenetic relationships of questionable tetraploid species Roegneria alashanica Keng, Roegneria magnicaespes (D.F. Cui) L.B. Cai, Roegneria elytrigioides C. Yen et J.L. Yang, Roegneria grandis Keng and Pseudoroegneria geniculata (Trin.) Á. Löve, the single copy sequences of the plastid acetyl-CoA carboxylase gene (Acc1) were analyzed among the five species and the related diploid and tetraploid species. The results indicated that: (a) R. alashanica contained one set of modified St genome which was closely related to the E^e genome, and the other set of genome was closely related to the P genome; (b) R. magnicaespes contained one set of St genome, the other set of genome might be closely related to the P genome. There are close affinities between R. magnicaespes and R. alashanica; (c) R. elytrigioides contained two sets of St genomes, and it is reasonable to be treated as Pseudoreogneria elytrigioides (C. Yen et J.L. Yang) B.R. Lu; (d) the genome of R. grandis should be designed as St^gY. The St^g genome was a differentiated form of the St genome in Pseudoroegneria and was homoeologous with the Y genome in Roegneria; (e) the genomic constitution of P. geniculata was similar to that of R. magnicaespes and R. alashanica and distinctly related to P. geniculata ssp. scythica (E^eSt). They should be treated as different species in different genera; and (f) the Y genome was possibly originated from the St genome, and was sister to the St, E^e, E^b and W genomes.     

Different responses to shade of evergreen and deciduous oak seedlings and the effect of acorn size

An evergreen oak species, Cyclobalanopsis multinervis, and a deciduous oak species, Quercus aliena var. acuteserrata were grown from acorns under two light levels (full sunlight and shade at about 18 % of full sunlight, simulating the light intensities in forest clearings and gaps, respectively) for one growing season. Three hypotheses were tested: (i) the deciduous species grows faster than the evergreen species in forest gaps and clearings; (ii) the deciduous species responds more strongly in terms of growth and morphology to variation in light climate than the evergreen species; and (iii) seedling size is positively correlated to acorn size. The results showed: (i) at both light levels, the deciduous seedlings gained significantly more growth in biomass and height than the evergreen seedlings; (ii) both species produced significantly more biomass in full sunlight than in shade, without showing any significant difference in height between treatments. Increase in light intensity improved the growth of the deciduous seedlings more strongly; (iii) at a similar age, the deciduous seedlings showed a greater response in leaf morphology and biomass allocation to variation in light levels, but when compared at a similar size, biomass allocation patterns did not differ significantly between species; (iv) bigger acorns tended to produce larger seedlings, larger leaf sizes and more leaf area, between and within species. These differences demonstrate that the deciduous species is gap-dependent and has the advantage over the evergreen species in forest gaps and clearings.     

Meiotic pairing behaviour reveals differences in genomic constitution between Hystrix patula and other species of the genus Hystrix Moench (Poaceae, Triticeae)

Interspecific and intergeneric hybridizations were carried out to evaluate the genomic relationships among species of Hystrix Moench and to study the relationships between Hystrix species and Psathyrostachys huashanica Keng (2n=2x=14, Ns^h). Meiotic pairing in hybrids of Hystrix duthiei ssp. duthiei × P. huashanica (2n=3x=21), Hystrix duthiei ssp. longearistata × P. huashanica (2n=3x=21) and H. patula × P. huashanica (2n=3x=21) averaged 5.18, 5.11 and 0.29 bivalents per cell, while H. patula × H. duthiei ssp. longearistata (2n=4x=28) averaged 25.36 univalents and 1.32 bivalents per cell, respectively. The results indicate that (i) H. duthiei ssp. duthiei and H. duthiei ssp. longearistata have one set of Ns genome from Psathyrostachys; (ii) H. patula has no Ns genome; (iii) genomes of H. duthiei ssp. duthiei and H. duthiei ssp. longearistata are non-homologous to those of H. patula. The genomic relationships between H. patula and other Hystrix species are also discussed.     

Meiotic studies ofElymus nutans andE. jacquemontii (Poaceae,Triticeae) and their hybrids withPseudoroegneria spicata and seventeenElymus species

Hybridizations ofElymus nutans andE. jacquemontii were carried out with one species ofPseudoroegneria (S genome), and 20Elymus species, each containing either of the SH, SY, SYH, or SYW genomes. Chromosome configurations were analysed at metaphase I of the two target taxa and their interspecific hybrids. It is concluded that (i)E. nutans is an allohexaploid containing the SYH genomes, andE. jacquemontii is an allotetraploid having the SY genomes; (ii) the genomic affinity is associated with the geographic distance between the species studied; (iii) minor genomic structural rearrangements have occurred within the hexaploid taxon ofE. nutans.     

Phylogenetic relationships and the maternal donor of Roegneria (Triticeae: Poaceae) based on three nuclear DNA sequences (ITS,Acc1, and Pgk1) and one chloroplast region (trnL-F)

Roegneria is a polyploid perennial genus in the tribe Triticeae. Some species of Roegneria are morphologically similar to genus Elymus and have been classified in Elymus. To investigate the delimitation and phylogenetic relationships of Roegneria, nuclear (ITS, Acc1, and Pgk1) and chloroplast (trnL–trnF) DNA regions were sequenced for 38 allopolyploid species and 32 diploid species of Triticeae. Phylogenetic analyses of nuclear DNA revealed that all Roegneria species were included in the St and Y genome clades, and that the Y genome was closely related to the V and Xp genomes. The chloroplast DNA dataset showed that Roegneria species were grouped with Pseudoroegneria species. The Pseudoroegneria species from the Middle East (P. libanotica and P. tauri) and Central Asia (P. strigosa) were more closely related to Roegneria species. The results suggested that: (i) the species containing the St and Y genomes should be segregated from Elymus and treated as a distinct genus, Roegneria, based on the genomic constitution; (ii) P. libanotica, P. tauri, and/or P. strigosa potentially served as the maternal donor of the St genome in Roegneria; (iii) The Y genome of Roegneria originated from a diploid Y genome species, and the V and Xp genomes may have contributed to Y genome formation; (iv) among Roegneria species of previously uncertain genomic constitution, R. seriotina was tetraploid and possessed the StY genomes, E. calcicolus was hexaploid with the StYH genomic constitution and should be classified in Campeiostachys, R. glaucifolia possessed the StStY genomes, and R. tschimganica had the genomic constitution St₁St₂Y.