Endo-allopolyploidy of autopolyploids and recurrent hybridization—A possible mechanism to explain the unresolved Y-genome donor in polyploid Elymus species (Triticeae: Poaceae)

The genus Elymus L. in the tribe Triticeae (Poaceae) includes economically and ecologically important forage grasses. The genus contains the pivotal St genome from Pseudoroegneria in combination with other genomes in the tribe. Many Elymus species are tetraploids containing the StY genomes. It is thought that polyploidization characterizes the speciation of the genus in which the Y is considered as another key genome. Based on data from cytological, genome in situ hybridization, and molecular studies, we hypothesized an endo-allopolyploidy origin of the StY-genome species from the autotetraploid Pseudoroegneria species. To test this hypothesis, we amplified, cloned, and sequenced five single-copy nuclear genes (i.e., alcohol dehydrogenase 1–3, Adh1–Adh3, RNA polymerase II, Rpb2; and Waxy) from Elymus, Pseudoroegneria, and Hordeum species. The phylogenetic trees constructed based on the sequencing analyses of all genes indicated that diploid and autotetraploid Pseudoroegneria species were closely related, although with considerable genetic variation in tetraploids. In addition, the StY-genome Elymus species tended to have a close relationship with the diploid and autotetraploid Pseudoroegneria species, although different phylogenetic relationships among the gene trees were detected. These results indicated that the StY-genome species may have an autotetraploid origin and experienced recurrent hybridization. The complex St genomes in Pseudoroegneria in the polyploid state may gain more opportunities for within-species differentiation and recurrent hybridization. As a result, series modified versions of St genomes evolved into the StY genomes in some Elymus species.     

青海高原披碱草属种间天然杂种的细胞学鉴定

利用重复序列探针染色体荧光原位杂交（FISH）和基因组原位杂交（GISH）技术,对采自青海高原披碱草属种间天然杂种进行细胞学鉴定,同时结合物种分布及形态学特征,共揭示6种不同天然杂种的类型。第一类为垂穗披碱草（Elymus nutans Griseb.）和鹅观草属（Roegneria C.Koch）物种间的天然杂种,染色体数为35,染色体组成为StStYYH;第二类为垂穗披碱草和达乌力披碱草种（Elymus dahuricus Turcz.ex Griseb.）间杂种,染色体数为42,染色体组成为StStHHYY;第三类为达乌力披碱草和老芒麦（Elymus sibiricus L.）种间杂种,染色体数为35,染色体组成为StStHHY;第四类为垂穗披碱草和糙毛以礼草（Kengilia hirsuta Keng）种间杂种,染色体数为42,染色体组成为StStYYHP;第五类为垂穗披碱草和大颖草（Kengilia grandiglumis Keng）种间杂种,染色体数为42,染色体组成为StStYYHP;第六类为糙毛以礼草和赖草（Leymus secalinus（Georgi） Tzvel.）种间杂种,染色体数为35,染色体组成为StYPNsXm。研究结果为进一步研究披碱草属种间杂交渐渗提供了重要参考资料;同时鉴定出的天然杂种可以作为潜在的种质资源在牧草或生态草育种中加以利用。     

Ubiquity of the St chloroplast genome in St-containing Triticeae polyploids.

Interspecific hybridization occurs between Tritceae species in the grass family (Poaceae) giving rise to allopolyploid species. To examine bias in cytoplasmic DNA inheritance in these hybridizations, the sequence of the 3' end of the chloroplast ndhF gene was compared among 29 allopolyploid Triticeae species containing the St nuclear genome in combination with the H, I, Ns, P, W, Y, and Xm nuclear genomes. These ndhF sequences were also compared with those from diploid or allotetraploid Triticeae species having the H, I, Ns, P, W, St, and Xm genomes. The cpDNA sequences were highly similar among diploid, allotetraploid, allohexaploid, and allooctoploid Triticeae accessions containing the St nuclear genome, with 0-6-nucleotide (nt) substitutions (0-0.8%) occurring between pairs of species. Neighbor-joining analysis of the sequences showed that the ndhF DNA sequences from species containing the St nuclear genome formed a strongly supported clade. The data indicated a strong preference for cpDNA inheritance from the St nuclear genome-containing parent in hybridizations between Triticeae species. This preference was independent of the presence of the H, I, Ns, P, W, and Xm nuclear genomes, the geographic distribution of the species, and the mode of reproduction. The data suggests that hybridizations having the St-containing parent as the female may be more successful.     

Evolutionary dynamics of the Pgk1 gene in the polyploid genus Kengyilia (Triticeae: Poaceae) and its diploid relatives

The level and pattern of nucleotide variation in duplicate gene provide important information on the evolutionary history of polyploids and divergent process between homoeologous loci within lineages. Kengyilia is a group of allohexaploid species with the StYP genomic constitutions in the wheat tribe. To investigate the evolutionary dynamics of the Pgk1 gene in Kengyilia and its diploid relatives, three copies of Pgk1 homoeologues were isolated from all sampled hexaploid Kengyilia species and analyzed with the Pgk1 sequences from 47 diploid taxa representing 18 basic genomes in Triticeae. Sequence diversity patterns and genealogical analysis suggested that (1) Kengyilia species from the Central Asia and the Qinghai-Tibetan plateau have independent origins with geographically differentiated P genome donors and diverged levels of nucleotide diversity at Pgk1 locus; (2) a relatively long-time sweep event has allowed the Pgk1 gene within Agropyron to adapt to cold climate triggered by the recent uplifts of the Qinghai-Tibetan Plateau; (3) sweep event and population expansion might result in the difference in the d(N)/d(S) value of the Pgk1 gene in allopatric Agropyron populations, and this difference may be genetically transmitted to Kengyilia lineages via independent polyploidization events; (4) an 83 bp MITE element insertion has shaped the Pgk1 loci in the P genome lineage with different geographical regions; (5) the St and P genomes in Kengyilia were donated by Pseudoroegneria and Agropyron, respectively, and the Y genome is closely related to the Xp genome of Peridictyon sanctum. The interplay of evolutionary forces involving diverged natural selection, population expansion, and transposable events in geographically differentiated P genome donors could attribute to geographical differentiation of Kengyilia species via independent origins.     

喜马红景天叶绿体基因组特征及其系统发育分析

以青藏高原特有药用植物——喜马红景天（Rhodiola himalensis）为试验材料,利用高通量测序技术对喜马红景天进行叶绿体基因组测序、组装和注释,获得完整的叶绿体基因组。结果显示：喜马红景天叶绿体基因组全长为151 074 bp,GC含量为37.8%,具有1个长单拷贝区、1个短单拷贝区和1对反向重复区的典型四分体结构,其序列长度分别为82 309、17 017、25 874 bp;叶绿体基因组共编码130个基因,其中编码蛋白的基因86个、编码tRNA的基因37个、编码rRNA的基因7个;叶绿体基因组共检测出25 513个密码子,其中编码亮氨酸（Leu）的密码子占比最大;喜马红景天IRa和IRb区的rps19和ycf1基因缺失,长单拷贝区的trnH基因收缩;喜马红景天与圣地红景天（R. sacra）亲缘关系最近;短单拷贝区域的单核苷酸多态性（SNP）变异频率最高。本研究报道了喜马红景天的叶绿体基因组,并对其进行了组装、注释和序列分析,为今后开展喜马红景天的遗传多样性研究和合理开发利用提供理论依据。     

Genome origin and phylogenetic relationships of Elymus villosus (Triticeae: Poaceae) based on single-copy nuclear Acc1, Pgk1, DMC1 and chloroplast trnL-F sequences

To investigate the genome origin and phylogenetic relationships of Elymus villosus, three single-copy nuclear gene (Acc1, Pgk1 and DMC1) and chloroplast trnL-F gene sequences of two accessions of E. villosus were analyzed with those of eighteen allotetraploids (StH, StY, StP and StE^e genomes) and thirty-five diploid taxa representing eighteen basic genomes in Triticeae. The results revealed that: (1) the genomic constitution of E. villosus is StH as Elymus; (2) North America Pseudoroegneria species served as the maternal donor during the allotetraploid speciation of E. villosus; (3) E. villosus is closely related to North America Elymus species; (4) it is reasonable to recognize the E. villosus as Elymus L. sensu stricto.     

Comparative phylogenetic analyses of Chinese Horsfieldia (Myristicaceae) using complete chloroplast genome sequences

The biologist's ruler for biodiversity is the species; accurate species identification is fundamental to the conservation of endangered species and in-depth biological scientific exploration. However, the delimitation and affinities of Horsfieldia in China has been controversial, owing in part to very low levels of molecular divergence within the family Myristicaceae. Because species boundaries and phylogenetic relationships within Horsfieldia are also unclear, 13 samples were collected across its distribution in China and their genomes were subjected to shotgun sequencing using Illumina platforms. A total of 40 487 994–84 801 416 pair-end clean reads were obtained and, after assembly, the complete chloroplast genome was recovered for all samples. Annotation analysis revealed a total of 112 genes, including 78 protein-coding genes, 30 transfer RNA, and 4 ribosomal RNA genes. Six variable loci (petN-psbM, trnH-psbA, ndhC-trnV, psbJ-psbL, ndhF, and rrn5-rrn23) were identified. Phylogenetic analyses strongly support the presence of four distinct species of Horsfieldia in China. In addition, samples that had been identified previously as Horsfieldia kingii (Hook. f.) Warb. were indistinguishable from those of H. prainii (King) Warb., suggesting that if H. kingii does occur in China, it was not collected in this study. Similarly, the chloroplast genome of one H. hainanensis Merr. sample from Guangxi province was identical to H. tetratepala C. Y. Wu, suggesting that the distribution range of H. hainanensis might be narrower than assumed previously. The phylogenetic relationships between the Chinese Horsfieldia species based on the whole chloroplast genomes was supported strongly, indicating the potential for using entire chloroplast genomes as super-barcodes for further resolution of the phylogeny of the genus Horsfieldia.     

Intergenomic rearrangements after polyploidization of Kengyilia thoroldiana (Poaceae: Triticeae) affected by environmental factors

Polyploidization is a major evolutionary process. Approximately 70–75% species of Triticeae (Poaceae) are polyploids, involving 23 genomes. To investigate intergenomic rearrangements after polyploidization of Triticeae species and to determine the effects of environmental factors on them, nine populations of a typical polyploid Triticeae species, Kengyilia thoroldiana (Keng) J.L.Yang et al. (2n = 6x = 42, StStPPYY), collected from different environments, were studied using genome in situ hybridization (GISH). We found that intergenomic rearrangements occurred between the relatively large P genome and the small genomes, St (8.15%) and Y (22.22%), in polyploid species via various types of translocations compared to their diploid progenitors. However, no translocation was found between the relatively small St and Y chromosomes. Environmental factors may affect rearrangements among the three genomes. Chromosome translocations were significantly more frequent in populations from cold alpine and grassland environments than in populations from valley and lake-basin habitats (P<0.05). The relationship between types of chromosome translocations and altitude was significant (r = 0.809, P<0.01). Intergenomic rearrangements associated with environmental factors and genetic differentiation of a single basic genome should be considered as equally important genetic processes during species'' ecotype evolution.     

Phylogenetic analysis of North American Elymus and the monogenomic Triticeae (Poaceae) using three chloroplast DNA data sets.

Although the monogenomic genera of the Triticeae have been analyzed in numerous biosystematic studies, the allopolyploid genera have not been as extensively studied within a phylogenetic framework. We focus on North American species of Elymus, which, under the current genomic system of classification, are almost all allotetraploid, combining the St genome of Pseudoroegneria with the H genome of Hordeum. We analyze new and previously published chloroplast DNA data from Elymus and from most of the monogenomic genera of the Triticeae in an attempt to identify the maternal genome donor of Elymus. We also present a cpDNA phylogeny for the monogenomic genera that includes more data than, and thus builds on, those previously published. The chloroplast DNA data indicate that Pseudoroegneria is the maternal genome donor to all but one of the Elymus individuals. There is little divergence among the Elymus and Pseudoroegneria chloroplast genomes, and as a group, they show little divergence from the rest of the Triticeae. Within the monogenomic Triticeae, the problematic group Thinopyrum is resolved as monophyletic on the chloroplast DNA tree. At the intergeneric level, the data reveal several deeper-level relationships that were not resolved by previous cpDNA trees.     

梅花草属叶绿体基因组进化分析

叶绿体基因组序列变异和基因组成等特征可有效反映植物类群间的系统发育和进化关系。本研究利用Illumina高通量测序平台对梅花草属（Parnassia）及其近缘属5种植物的叶绿体基因组进行测序和组装,同时基于已发表的近缘种叶绿体基因组信息,对梅花草属叶绿体基因组结构特征、序列遗传变异和蛋白编码基因密码子偏好性比对分析。结果显示：梅花草属叶绿体基因组整体结构较为保守,均为四分体结构;梅花草多个基因出现假基因化,而本属其他物种叶绿体基因组成一致,均编码115个基因;与近缘属物种相比,本属所有物种均丢失rpl16基因的内含子;蛋白质编码基因的非同义/同义替代率比值较低,叶绿体基因可能经历纯化选择作用;密码子偏好性聚类结果与蛋白编码序列重建的系统发育关系结果一致。本研究表明选择压力可能在梅花草属叶绿体基因组蛋白编码基因进化过程中发挥作用,有助于进一步理解梅花草属植物的进化和适应机制。     

兰州市南北两山5种典型人工林凋落物的水文功能

结合野外实地观测和室内浸水试验,对兰州市南北两山5种典型人工林(侧柏林、刺槐林、新疆杨林、侧柏+刺槐混交林、新疆杨+刺槐混交林)凋落物的持水特性进行研究。结果表明: 不同林分类型凋落物的蓄积量在13.50～47.01 t·hm^-2,依次为新疆杨+刺槐混交林＞侧柏+刺槐混交林＞侧柏林＞刺槐林＞新疆杨林。除侧柏林外,其他林分类型的半分解层蓄积量所占比例均高于未分解层。凋落物最大持水率在190.8%～262.7%,新疆杨+刺槐混交林最大,侧柏林最小。凋落物最大持水量在35.29～123.59 t·hm^-2,依次为新疆杨+刺槐混交林＞侧柏+刺槐混交林＞刺槐林＞侧柏林＞新疆杨林。凋落物的吸水速率在浸水0～1 h直线下降,1 h后降幅减缓,未分解层凋落物吸水速率小于半分解层。最大拦截量和有效拦截量(深)均为新疆杨+刺槐混交林＞侧柏+刺槐混交林＞侧柏林＞刺槐林＞新疆杨林,且新疆杨+刺槐混交林有较高的有效拦截率,表明新疆杨+刺槐混交林在兰州市南北两山有较好的保持水土和涵养水源能力。     

Phylogenetic relationships in Elymus (Poaceae: Triticeae) based on the nuclear ribosomal internal transcribed spacer and chloroplast trnL-F sequences

To estimate the phylogenetic relationship of polyploid Elymus in Triticeae, nuclear ribosomal internal transcribed spacer (ITS) and chloroplast trnL-F sequences of 45 Elymus accessions containing various genomes were analysed with those of five Pseudoroegneria (St), two Hordeum (H), three Agropyron (P) and two Australopyrum (W) accessions. The ITS sequences revealed a close phylogenetic relationship between the polyploid Elymus and species from the other genera. The ITS and trnL-F trees indicated considerable differentiation of the StY genome species. The trnL-F sequences revealed an especially close relationship of Pseudoroegneria to all Elymus species included. Both the ITS and trnL-F trees suggested multiple origins and recurrent hybridization of Elymus species. The results suggested that: the St, H, P, and W genomes in polyploid Elymus were donated by Pseudoroegneria, Hordeum, Agropyron and Australopyrum, respectively, and the St and Y genomes may have originated from the same ancestor; Pseudoroegneria was the maternal donor of the polyploid Elymus; and some Elymus species showed multiple origin and experienced recurrent hybridization.     

Comparison of Acetyl-CoA carboxylase 1 (Acc-1) gene diversity among different Triticeae genomes

It has widely been documented that life form and mating system have significant influences on genetic diversity. In the tribe Triticeae, several genera contain both annual and perennial species, whereas other genera comprise strictly annual or perennial species. It was suggested that Triticeae annuals have originated from Triticeae perennials. The present study aims to analyze nucleotide diversity of Acc-1 gene among different Triticeae genomes, and attempts to link effects of life history (annuals and perennials) and mating systems. The nucleotide diversity of 364 Acc-1 sequences in Triticeae species was characterized. The highest estimates of nucleotide diversity values (π = 0.01919, θ = 0.03515) were found for the Ns genome among the genomes analyzed. Nucleotide diversities in the D genome and Ns genome of polyploids are higher than those in respective genomes of diploids, while in the St genome of polyploids, it is lower than that in the St genome of diploids. The averaged π value (0.013705) in the genomes of perennials is more than twice of the value (0.00508) in the genomes of annuals. The averaged π value (0.01323) in the genomes of outcrossing species is two-fold of the value (0.005664) in the genomes of selfer. Our results suggested that the evolutionary history and mating system may play an important role in determining nucleotide diversity of Acc-1 gene in each genome.     

Molecular phylogeny revealed complex evolutionary process in Elymus species

Recent molecular phylogenetic studies on Elymus have added to our understanding of the origination of Elymus species. However, evolutionary dynamics and speciation of most species in Elymus are unclear. Molecular phylogeny has demonstrated that reticulate evolution has occurred extensively in the genus, as an example, the largest subunit of RNA polymerase II (rpb2) and phosphoenolpyruvate carboxylase (pepC) data revealed two versions of the St genome, St1 and St2contributing to speciation of E. caninus. Phylogenetic analyses of E. pendulinus uncovered additional genome-level complexity. Our data indicated that both chloroplast and nuclear gene introgression have occurred in the evolutionary process of E. pendulinus. Non-donor species genomes have been detected in severalElymus species, such as in allohexaploid E. repens (StStStStHH), a Taeniatherum-like (Ta genome in Triticeae) GBSSI sequence, Bromus- (Bromeae) and Panicum-like (Paniceae) ITS sequences have been detected. The chloroplast DNA data indicated that Pseudoroegneria is the maternal genome donor to Elymus species, but whether different Elymus species originated from different St donors remains an open question. The origin of the Y genome in Elymus is puzzling. It is clear that the Ygenome is distinct from the St genome, but unclear on the relationships of Y to other genomes in Triticeae. Introgressive hybridization may be an important factor complicating the evolutionary history of the species in Elymus. The extent of introgression and its role in creating diversity in Elymus species should be the objective of further investigations.     

Genetic Relationships Among Five Basic Genomes St,E,A,B and D in Triticeae Revealed by Genomic Southern and in situ Hybridization

The St and E are two important basic genomes in the perennial tribe Triticeae (Poaceae). They exist in many perennialspecies and are very closely related to the A, B and D genomes of bread wheat (Triticum aestivum L.). Genomic Southernhybridization and genomic in situ hybridization (GISH) were used to analyze the genomic relationships between the twogenomes (St and E) and the three basic genomes (A, B and D) of T. aestivum. The semi-quantitative analysis of the Southernhybridization suggested that both St and E genomes are most closely related to the D genome, then the A genome, andrelatively distant to the B genome. GISH analysis using St and E genomic DNA as probes further confirmed the conclusion.St and E are the two basic genomes of Thinopyrum ponticum (StStE~eE~bE~x) and Th. intermedium (StE~eE~b), two perennialspecies successfully used in wheat improvement. Therefore, this paper provides a possible answer as to why most of thespontaneous wheat-Thinopyrum translocations and substitutions usually happen in the D genome, some in the A genomeand rarely in the B genome. This would develop further use of alien species for wheat improvement, especially thosecontaining St or E in their genome components.     

Genome discrimination by in situ hybridization in Icelandic species of Elymus and Elytrigia (Poaceae: Triticeae).

The genome constitution of Icelandic Elymus caninus, E. alaskanus, and Elytrigia repens was examined by fluorescence in situ hybridization using genomic DNA and selected cloned sequences as probes. Genomic in situ hybridization (GISH) of Hordeum brachyantherum ssp. californicum (diploid, H genome) probe confirmed the presence of an H genome in the two tetraploid Elymus species and identified its presence in the hexaploid Elytrigia repens. The H chromosomes were painted uniformly except for some chromosomes of Elytrigia repens which showed extended unlabelled pericentromeric and subterminal regions. A mixture of genomic DNA from H. marinum ssp. marinum (diploid, Xa genome) and H. murinum ssp. leporinum (tetraploid, Xu genome) did not hybridize to chromosomes of the Elymus species or Elytrigia repens, confirming that these genomes were different from the H genome. The St genomic probe from Pseudoroegneria spicata (diploid) did not discriminate between the genomes of the Elymus species, whereas it produced dispersed and spotty hybridization signals most likely on the two St genomes of Elytrigia repens. Chromosomes of the two genera Elymus and Elytrigia showed different patterns of hybridization with clones pTa71 and pAes41, while clones pTa1 and pSc119.2 hybridized only to Elytrigia chromosomes. Based on FISH with these genomic and cloned probes, the two Elymus species are genomically similar, but they are evidently different from Elytrigia repens. Therefore the genomes of Icelandic Elymus caninus and E. alaskanus remain as StH, whereas the genomes of Elytrigia repens are proposed as XXH.     

四种菊头蝠染色体组型分析

本文分析了皮氏菊头蝠(R. pearsoni chinensis),鲁氏菊头蝠(R.rouxi sinicus),角菊头蝠(R.cornutus pumilus)及中菊头蝠(R.affinis)的常规核型,现报道如下。     

Molecular confirmation of the genomic constitution of <Emphasis Type="Italic">Douglasdeweya</Emphasis> (Triticeae: Poaceae): demonstration of the utility of the 5S rDNA sequence as a tool for haplome identification

A new genus Douglasdeweya containing the two species, Douglasdeweya deweyi and D. wangii was published in 2005 by Yen et al. based upon the results of cytogenetical and morphological findings. The genome constitution of Douglasdeweya-PPStSt-allowed its segregation from the genus Pseudoroegneria which contains the StSt or StStStSt genomes. Our previous work had demonstrated the utility of using 5S rDNA units, especially the non-transcribed spacer sequence variation, for the resolution of genomes (haplomes) previously established by cytology. Here, we show that sequence analysis of the 5S DNA units from these species strongly supports the proposed species relationships of Yen et al. (Can J Bot 83:413-419, 2005), i.e., the PP genome from Agropyron and the StSt genome from Pseudoroegneria. Analysis of the 5S rDNA units constitutes a powerful tool for genomic research especially in the Triticeae.     

Genetic Relationships Among Five Basic Genomes St, E, A, B and D in Triticeae Revealed by Genomic Southern and in situ Hybridization

The St and E are two important basic genomes in the perennial tribe Triticeae （Poaceae）. They exist in many perennial species and are very closely related to the A, B and D genomes of bread wheat （Triticum aestivum L.）. Genomic Southern hybridization and genomic in situ hybridization （GISH） were used to analyze the genomic relationships between the two genomes （St and E） and the three basic genomes （A, B and D） of T. aestivum. The semi-quantitative analysis of the Southern hybridization suggested that both St and E genomes are most closely related to the D genome, then the A genome, and relatively distant to the B genome. GISH analysis using St and E genomic DNA as probes further confirmed the conclusion. St and E are the two basic genomes of Thinopyrum ponticum （StStE^eE^bE^x） and Th. intermedium （StE^eE^b）, two perennial species successfully used in wheat improvement. Therefore, this paper provides a possible answer as to why most of the spontaneous wheat-Thinopyrum translocations and substitutions usually happen in the D genome, some in the A genome and rarely in the B genome. This would develop further use of alien species for wheat improvement, especially those containing St or E in their genome components.     

Analysis of phylogenetic relationships of Potamogeton species in China based on chloroplast trnT-trnF sequences

Sequences of the chloroplast trnT-trnF region were analyzed for species of the genus Potamogeton distributed in China to reconstruct phylogenetic relationships. The phylogenetic analyses showed that the genus Potamogeton could be divided into two clades. Eighteen species formed a monophyletic clade while the remaining four formed a second, distinguishable one, supporting the conventional treatment that the genus Potamogeton contains the submerged linear-leaved group and the submerged broad-leaved group. The first clade, which represented the subgenus Potamogeton, could be further divided into two subclades. The second clade, which represented the subgenus Coleogeton, displayed a close phylogenetic relationship with the subgenus Potamogeton and occupied a unique position within the genus Potamogeton. This finding suggested that the treatment of the subgenus Coleogeton, which was once regarded as the genus Stuckenia Börner, may need to be reconsidered. Furthermore, identification of maternal donors of some hybrids was successfully applied based on sequence of maternally inherited chloroplast genome. The female parents of three putative hybrids, P. × malainoides, P. × anguillanus, and P. × orientalis, were proved to be accordant with previous morphological conclusions.