EVOLUTION OF POPULUS NIGRA (SECT. AIGEIROS):INTROGRESSIVE HYBRIDIZATION AND THE CHLOROPLAST CONTRIBUTION OF POPULUS ALBA (SECT. POPULUS)

Restriction site variation in chloroplast DNA and nuclear ribosomal DNA was examined in 16 accessions from the Salicaceae comprising ten species of Populus and one outgroup species of Salix. Forty-nine restriction site mutations in the chloroplast DNAs were used to generate one most parsimonious phylogenetic tree. This tree indicates that all varieties of P. nigra (black poplars of sect. Aigeiros) have a chloroplast genome, maternally inherited, derived from the clade including the white poplars (P. alba and segregate species of sect. Populus) and divergent from the American cottonwoods of their own section. Twenty-one restriction site mutations in the nuclear ribosomal DNAs generated a single most parsimonious phylogenetic tree that indicates that the nuclear genome ofP. nigra is distinct from both the white poplars and American cottonwoods. The incongruity of these independent molecular phylogenies provides evidence for an unusual origin of the black poplars. Populus alba or its immediate ancestor acted as the maternal parent in a hybridization event with the paternal lineage of P. nigra. Subsequent backcrosses to the paternal species gave rise to the extant P. nigra with a chloroplast genome of P. alba and the nuclear genome of the paternal species. These hybridization and introgression events must have pre-dated the divergence of the black poplar varieties. The biphyletic nature of the P. nigra genomes suggests that dependency on one class of molecular or morphological markers or the merging of the two kinds of data sets to derive accurate estimates of true phylogenies could be misleading in plants.     

Comparative genome mapping among Populus adenopoda, P. alba, P. deltoides, P. euramericana and P. trichocarpa

Among the genus Populus, the sections Populus (white poplar), Aigeiros Duby (black poplar) and Tacamahaca Spach contain many tree species of economical and ecological important properties. Two parental maps for the inter-specific hybrid population of Populus adenopoda × P. alba (two species of Populus section) were constructed based on SSR and SRAP markers by means of a two-way pseudo-test cross mapping strategy. The same set of SSR markers developed from the P. trichocarpa (belonging to Tacamahaca section) genome which were used to construct the maps of P. deltoides and P. euramericana (two species of Aigeiros section) was chosen to analyze the genotype of the experimental population of P. adenopoda × P. alba. Using the mapped SSR markers as allelic bridges, the alignment of the white and black poplar maps to each other and to the P. trichocarpa physical map was conducted. The alignment showed high degree of marker synteny and colinearity and the closer relationship between Aigeiros and Tacamahaca sections than that of Populus and Tacamahaca. Moreover, there was evidence for the chromosomal duplication and inter-chromosomal reorganization involving some poplar linkage groups, suggesting a complicated course of fission or fusion in one of the lineages. A poplar consensus map based on the comparisons could be constructed will be useful in practical applications including marker assisted selection.     

衣藻属的系统发育分析——基于形态形状和nrDNA ITS序列

通过实验分析莱茵衣藻 ( Chlamydomonas reinhardtii) 1个种和互连网获得衣藻属 1 5个种及丝藻属 1个种 ( Ulothrix zonata) ,共 1 7个种的 nr DNA ITS序列 ,并以 U.zonata为外类群 ,采用计算机分析软件包对其进行分析及构建分子系统发育树图。同时以 1 2个传统分类性状 ,对此 1 6种衣藻构建数据矩阵 ;以 U.zonata动孢子的相应性状为外类群原始性状 ,用Wagner法在计算机上对其进行分枝分析 ;然后比较并分析分子系统树和表征性状分支分析树的异同。初步尝试以 ITS分子序列系统发育分析作为传统性状分析的补充来研究衣藻种间的亲缘关系。     

The phylogeny of Schistidium (Bryophyta, Grimmiaceae) based on primary and secondary structure study of nuclear rDNA internal transcribed spacers

The phylogeny of Schistidium (Bryophyta, Grimmiaceae) was studied on the basis of nucleotide sequences of internal transcribed spacers ITS1-2 of nuclear DNA and trnT-trnD region of chloroplast DNA. The consistency of phylogenetic trees constructed from nuclear and chloroplast sequences was shown. A basal grade and two large clades were resolved on the phylogenetic trees. Morphological characteristics specific for these clades were described. ITS1 and ITS2 secondary structures of Schistidium species were modeled using thermodynamic criteria. Four different structures of the longest ITS1 hairpin were identified. Possible paths of Schistidium evolution were considered based on the four types of ITS1 secondary structure and phylogenetic trees.     

Restriction fragment length polymorphisms of a chloroplast photosystem II gene from poplar and their use for species identification.

In a total DNA library from the poplar clone Beaupré (Populus trichocarpa x P. deltoides) one DNA clone was found to identify restriction site polymorphisms in different poplar species. This clone represents a cpDNA gene that shows close homology to a photosystem II gene of pea and spinach coding for the D2 protein and the 44 kDa reaction centre. In Southern blot analysis this probe identified interspecific restriction site variation among the different poplar species; intraspecific variation was not detectable. As the chloroplast genome is maternally inherited in poplars this cpDNA probe was used for identification of P. nigra or P. deltoides as the seed parents of F1 hybrid trees in natural stands of western Germany.     

Causes of Size Homoplasy Among Chloroplast Microsatellites in Closely Related <Emphasis Type="Italic">Clusia</Emphasis> Species

Chloroplast DNA sequences and microsatellites are useful tools for phylogenetic as well as population genetic analyses of plants. Chloroplast microsatellites tend to be less variable than nuclear microsatellites and therefore they may not be as powerful as nuclear microsatellites for within-species population analysis. However, chloroplast microsatellites may be useful for phylogenetic analysis between closely related taxa when more conventional loci, such as ITS or chloroplast sequence data, are not variable enough to resolve phylogenetic relationships in all clades. To determine the limits of chloroplast microsatellites as tools in phylogenetic analyses, we need to understand their evolution. Thus, we examined and compared phylogenetic relationships of species within the genus Clusia, using both chloroplast sequence data and variation at seven chloroplast microsatellite loci. Neither ITS nor chloroplast sequences were variable enough to resolve relationships within some sections of the genus, yet chloroplast microsatellite loci were too variable to provide any useful phylogenetic information. Size homoplasy was apparent, caused by base substitutions within the microsatellite, base substitutions in the flanking regions, indels in the flanking regions, multiple microsatellites within a fragment, and forward/reverse mutations of repeat length resulting in microsatellites of identical base composition that were not identical by descent.     

Molecular phylogeny and evolution of Caricaceae based on rDNA internal transcribed spacers and chloroplast sequence data

This study focused on clarifying phylogenetic relationships and evolution within Caricaceae. Our phylogenetic analysis based on nucleotide sequences from the ITS of the ribosomal DNA and three chloroplast fragments (matK, trnL-trnF, and psbA-trnH) included 29 taxa belonging to five genera: the neotropical genera Carica, Vasconcellea, Jarilla, and Jacaratia and the equatorial African genus Cylicomorpha. Having a relatively low mutation rate, matK, and trnL-trnF were used for estimating relationships at the generic level, while intrageneric evolution within Vasconcellea was studied with the more variable ITS and psbA-trnH sequences. Gaps, coded as binary characters, were added to the sequence alignments before performing Maximum Parsimony and Maximum Likelihood analyses. Monophyly of Caricaceae as well as phylogenetic distance between Carica and Vasconcellea species, previously belonging to the same genus, and monophyly of the resurrected genus Vasconcellea were emphasized. Within Vasconcellea, the largest genus of this family, two well-confirmed evolutionary lineages could be discerned: (1) V.xheilbornii, V. weberbaueri, V. stipulata, and V. parviflora and (2) a clade holding all other taxa of the genus. Incongruence between nuclear ITS and chloroplast psbA-trnH datasets, shown to be significantly caused by some taxa of the genus Vasconcellea, indicated that reticulate events in this genus might be more frequent than previously suspected. Moreover, intra-individual ITS sequence heterogeneity provided further evidence for the hybrid or introgressed origin of different taxa and one presumed hybrid belonging to this genus.     

Study of genetic relationships and phylogeny of the native Populus in Southwest China based on nucleotide sequences of chloroplast trnT–trnF and nuclear DNA

Unique historical factors and ecological conditions make Southwest China a natural distribution and variation center for trees of the genus Populus in China. However, little is currently known about the native poplars occurring in this region, and considerable doubt still exists regarding the classification and evolutionary relationships of poplar species. In this study, nuclear and chloroplast DNA (cpDNA) sequences were utilized to determine the genetic relationships and phylogeny of Populus species in Southwest China. The results suggest that P. pseudoglauca belongs to the section of Tacamahaca. Further, P. schneideri may be a natural hybrid of P. kangdingensis and P. cathayana and, thus, it should likely not be regarded as a variety of P. kangdingensis, as in the existing classification system. In addition, cluster analyses showed that P. gonggaensis may be derived from a cross between species of section Leucoides and P. cathayana or P. schneideri of section Tacamahaca, although it is still doubtful whether P. gonggaensis can be regarded as a separate species, due to its narrow distribution range. The parents of the Luding poplar may be P. yunnanensis and P. lancifolia. P. butuoensis showed a close affinity to species of section Leucoides and had a close relationship with P. gonggaensis or P. lasiocarpa. However, further research is needed in order to appropriately classify these as species or varieties. The incongruence between phylogenetic trees based on nuclear- and chloroplast-DNA sequence data may be due to the different inheritance patterns between nuclear- and cpDNA genome.     

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.     

ITS1 Sequences of Nuclear Ribosomal DNA in Wild Rices and Cultivated Rices of China and Their Phylogenetic Implications

he first internal transcribed spacer (ITS1) of nuclear ribosomal DNA of three wild rice species and two subspecies of cultivated rice, which are distributed in China, was amplified using PCR technique and sequenced with automated fluorescent sequencing. The sequences of ITS1 ranged from 193 bp to 218 bp in size and G/C content varied from 69.3%to 72.7%. In pairwise comparison among the five taxa, sequence site divergence ranged from 1.5 % to 10.6%. Phylogenetic analysis of ITS1 sequences using Wagner parsimony generated a single well-resolved tree, which revealed that Oryza rufipogon was much more closely related to cultivated rice species than to the other two wild species. Oryza granulata was less closely related to either cultivated rice species or the other two wild species, and might be a unique and isolated taxon in the genus Oryza. The phylogenetic relationships of the three wild rice species and two cultivated rice subspecies inferred from ITS1 sequences is highly concordant with those based on the molecular evidence from isozyme, chloroplast DNA (cpDNA), mitochondrial DNA (mtDNA) and nuclear DNA (nDNA) of the genus Oryza.     

Wax plants disentangled: a phylogeny of Hoya (Marsdenieae, Apocynaceae) inferred from nuclear and chloroplast DNA sequences

Hoya (Marsdenieae, Apocynaceae) includes at least 200 species distributed from India to the Pacific Islands. We here infer major species groups in the genus based on combined sequences from the chloroplast atpB-rbcL spacer, the trnL region, and nuclear ribosomal DNA ITS region for 42 taxa of Hoya and close relatives. To assess levels of ITS polymorphism, ITS sequences for a third of the accessions were obtained by cloning. Most ITS clones grouped by species, indicating that speciation in Hoya usually predates ITS duplication. One ITS sequence of H. carnosa, however, grouped with a sequence of the morphologically similar H. pubicalyx, pointing to recent hybridization or the persistence of paralogous copies through a speciation event. The topology resulting from the combined chloroplast and nuclear data recovers some morphology-based sections, such as Acanthostemma and Eriostemma, as well as a well-supported Australian/New Guinean clade. The combined data also suggest that morphological adaptations for ant-symbiosis evolved at least three times within Hoya.     

Its evolution in Platanus (Platanaceae): homoeologues, pseudogenes and ancient hybridization

BACKGROUND AND AIMS: Platanaceae is an old family of angiosperms extending back to the Early Cretaceous but consisting of a single extant genus, Platanus. Species of Platanus have long been known to hybridize, and the London plane, Platanus x hispanica, is a well-known example of a hybrid species that formed in historical times. In addition, morphological studies have suggested past interspecific or interlineage hybridization and reticulation as possibly important factors in the evolution of the genus. This study aims at unravelling the complex evolutionary information contained in internal transcribed spacer (ITS) sequences. METHODS: The ITS of the 35S nuclear ribosomal DNA are biparentally inherited, multi-copy markers with a high potential for resolving intrageneric relationships especially when ancient hybridization (reticulation) is involved. Phylogenetic trees, splits graphs and motif analysis are used to extract phylogenetic information from 223 cloned ITS sequences, representing ten species and varieties of Platanus. Non-pseudogenous and pseudogenous sequence motives are assessed to explain how different evolutionary modes contribute to possibly conflicting character state patterns in the ITS. KEY RESULTS: It was found that putative non-functional ITS copies ('pseudogenes') form distinct groups in phylograms and splits graphs, and that pseudogenous lineages reflect ancient hybridization events conserved in the ITS. Specifically, pseudogenous clones of an 'Atlantic' North American clade share sequence motives with non-pseudogenous clones of the western ('Pacific') North American P. racemosa species aggregate. In addition, evidence was found for recent lateral gene flow as a possible factor in the evolution of the central Mexican P. rzedowskii. CONCLUSIONS: Broad ITS data sets that cover intra- and interindividual variability reveal past and ongoing speciation processes in Platanus. Evolutionary pathways can be visualized with splits graphs, but not with bifurcating trees.     

中国野生稻及栽培稻核糖体DNA第一转录间隔区序列分析及其系统学意义

用 PCR技术从产于我国的 3种野生稻和亚洲栽培稻的 2个亚种中特异地扩增和测序了 r DNA的第一转录间隔区。普通野生稻 (Oryza rufipogon)、药用野生稻 (O.officinalis)、疣粒野生稻 (O.granu-lata)和栽培稻的两个亚种 (O.sativa ssp.indica,O.sativa ssp.japonica)的 ITS1序列为 1 93bp、1 94bp、2 1 8bp、1 94bp和 1 94bp,它们的 G/ C含量为 69.3%～ 72 .7% ,序列中位点趋异率为 1 .5%～ 1 0 .6%。序列的相似性比较和简约性分支分析的结果表明 ,普通野生稻与栽培稻的两个亚种之间的亲缘关系最为密切 ;药用野生稻与普通野生稻和与栽培稻的两个亚种的相似性都为 82 % ,说明它与 AA基因组有一定的亲缘关系 ;疣粒野生稻与普通野生稻、药用野生稻和栽培稻两个亚种的亲缘关系相对较远 ,它在稻属中可能是一个系统地位较独特的类群。以 ITS1序列构建的 3种野生稻和 2个栽培稻亚种的系统发育关系与前人用同工酶、叶绿体 DNA、线粒体 DNA和核 DNA资料重建的稻属的系统发育关系基本一致     

Molecular evidence for a Mediterranean origin of the Macaronesian endemic genus Argyranthemum (Asteraceae)

The internal transcribed spacers (ITS) of nuclear ribosomal DNA were sequenced for 52 species from 32 genera and eight subtribes of Anthemideae. Phylogenetic analyses of ITS data generated trees that are largely incongruent with the recent classification of Anthemideae; most of the subtribes examined are not resolved as monophyletic. However, ITS trees are congruent with morphological, isozyme, phytochemical, and chloroplast DNA (cpDNA) restriction site data in supporting a Mediterranean origin for Argyranthemum, the largest endemic genus of the Atlantic oceanic islands. A combined analysis of ITS sequences and cpDNA restriction sites indicates that Argyranthemum is sister to the other three genera of Chrysantheminae (i.e., Chrysanthemum, Heteranthemis, and Ismelia). Times of divergence of Argyranthemum inferred from the ITS sequences ranged between 0.26 and 2.1 million years ago (mya) and are lower than values previously reported from isozyme and cpDNA data (1.5-3.0 mya). It is likely that rate heterogeneity of the ITS sequences in the Anthemideae accounts for the low divergence-time estimates. Comparison of data for 20 species in Argyranthemum and Chrysantheminae indicates that the cpDNA restriction site approach provided much more phylogenetic information than ITS sequences. Thus, restriction site analyses of the entire chloroplast genome remain a valuable approach for studying recently derived island plants.     

Phylogenetic Use of Noncoding Regions in the GenusGentianaL.: ChloroplasttrnL (UAA) Intron versus Nuclear Ribosomal Internal Transcribed Spacer Sequences

Sequence divergence was estimated within noncoding sequences of both chloroplast DNA (cpDNA)trnL (UAA) intron and nuclear ribosomal DNA (nrDNA) internal transcribed spacer sequences (ITS1 and ITS2) for 10 species of the genusGentianaL. (Gentianaceae). Comparisons of evolutionary rates among these sequences (cpDNA versus nrDNA, ITS1 versus ITS2) were performed. It appears that sequence divergence is on average two to three times higher in ITSs than in thetrnL intron sequences and higher in ITS1 than in ITS2. Both the cpDNA intron and ITSs of nrDNA give concordant phylogenetic trees. However, the ITS-based phylogeny displays higher bootstrap values. At the intrageneric level, at least inGentiana,ITSs (especially ITS2) sequences seem to be more appropriate in the assessment of plant phylogenies. Nevertheless, the cpDNAtrnL intron seems to be preferable at the intergeneric level.     

基于DNA非编码区序列探讨罗布麻的分类问题

通过测定中国产罗布红麻、大叶白麻和白麻的ITSt、rnL内含子和trnL-F非编码区序列,并与其它相关植物的相应序列比较,探讨“罗布麻”的系统分类。结果显示:罗布红麻、大叶白麻和白麻3种植物的ITS序列完全一致,而与罗布麻属加拿大麻的ITS序列相差较大;在trnL内含子和trnL-F非编码区,大叶白麻和白麻的完全一致,与罗布红麻之间仅有3个位点的变异,而加拿大麻与美国茶叶花在这3个位点却和白麻属的完全一致等。以上结果显示在ITSt、rnL内含子和trnL-F非编码区,白麻属和罗布麻属之间没有本质区别,罗布红麻与大叶白麻和白麻之间的亲缘关系可能较罗布红麻与罗布麻属内其它物种间的亲缘关系更近。遗传距离与系统树的分析结果进一步支持以上推论。建议撤销白麻属,将大叶白麻和白麻合并到罗布麻属。     

Study of the evolutionary relationships among Limonium species (Plumbaginaceae) using nuclear and cytoplasmic molecular markers

The genus Limonium, due to the patchiness of the natural habitats of its species as well as the high frequency of hybridization and polyploidy and the possibility of reproduction by apomixis, provides an example of all the principal mechanisms of rapid speciation of plants. As an initial study of evolution in this genus, we have analyzed intra- and interspecific variability in 17 species from section Limonium, the largest in the genus, based on RFLPs of cpDNA and nuclear rDNA ITS sequences. In the cpDNA analysis, 21 restriction enzymes were used, resulting in 779 fragments, 490 of which were variable and 339 parsimony informative. L. furfuraceum exhibited two relatively divergent cpDNA haplotypes. The relationships found among the species based on cpDNA restriction fragments were coincident using different methods of phylogenetic analysis. Due to the presumed reticulate evolution in the genus Limonium, the comparison of these results with data from the nuclear DNA was necessary; ITS sequences were analyzed. The final alignment contained 488 characters, of which 198 were variable and 156 parsimony informative. Two relatively divergent ITS types were present at the intraindividual level in L. delicatulum, a triploid species. Each type was related to ITS from different groups of diploid Limonium species, one with a base haploid chromosome number n = 8 (represented by L. cossonianum) and the other with n = 9 (represented by L. minutum). The different phylogenetic inference methods used for the analysis of ITS sequences rendered very similar topologies. In general, the relationships among the species studied were coincident with those obtained with the chloroplast genome. Both nuclear and cytoplasmic markers support the polyphyly of section Limonium, with at least two species, L. narbonense and L. vulgare, clearly divergent from the rest. Moreover, the remaining subsections into which section Limonium is currently divided seem to be artificial.     

DISCORDANCE BETWEEN NUCLEAR AND CHLOROPLAST PHYLOGENIES IN THE HEUCHERA GROUP (SAXIFRAGACEAE)

Various factors, including taxon density, sampling error, convergence, and heterogeneity of evolutionary rates, can potentially lead to incongruence between phylogenetic trees based on different genomes. Particularly at the generic level and below, chloroplast capture resulting from hybridization may distort organismal relationships in phylogenetic analyses based on the chloroplast genome, or genes included therein. However, the extent of such discord between chloroplast DNA (cpDNA) trees and those trees based on nuclear genes has rarely been assessed. We therefore used sequences of the internal transcribed spacer regions (ITS-1 and ITS-2) of nuclear ribosomal DNA (rDNA) to reconstruct phylogenetic relationships among members of the Heuchera group of genera (Saxifragaceae). The Heuchera group presents an important model for the analysis of chloroplast capture and its impact on phylogenetic reconstruction because hybridization is well documented within genera (e.g., Heuchera), and intergeneric hybrids involving six of the nine genera have been reported. An earlier study provided a well-resolved phylogenetic hypothesis for the Heuchera group based on cpDNA restriction-site variation. However, trees based on ITS sequences are discordant with the cpDNA-based tree. Evidence from both morphology and nuclear-encoded allozymes is consistent with the ITS trees, rather than the cpDNA tree, and several points of phylogenetic discord can clearly be attributed to chloroplast capture. Comparison of the organellar and ITS trees also raises the strong likelihood that ancient events of chloroplast capture occurred between lineages during the early diversification of the Heuchera group. Thus, despite the many advantages and widespread use of cpDNA data in phylogeny reconstruction, comparison of relationships based on cpDNA and ITS sequences for the Heuchera group underscores the need for caution in the use of organellar variation for retrieving phylogeny at lower taxonomic levels, particularly in groups noted for hybridization.     

Phylogenetic utility of the internal transcribed spacers of nuclear ribosomal DNA in plants: an example from the compositae.

The internal transcribed spacer (ITS) region of 18-26S nuclear ribosomal DNA was sequenced in 12 representatives of the Compositae subtribe Madiinae and two outgroup species to assess its utility for phylogeny reconstruction. High sequence alignability and minimal length variation among ITS 1, 5.8S, and ITS 2 sequences facilitated determination of positional homology of nucleotide sites. In pairwise comparisons among Madiinae DNAs, sequence divergence at unambiguously aligned sites ranged from 0.4 to 19.2% of nucleotides in ITS 1 and from 0 to 12.9% of nucleotides in ITS 2. Phylogenetic relationships among ITS sequences of Hawaiian silversword alliance species (Argyroxiphium, Dubautia, and Wilkesia) and California tarweed taxa in Adenothamnus, Madia, Raillardella, and Raillardiopsis are highly concordant with a chloroplast DNA-based phylogeny of this group. Maximally parsimonious trees from ITS and chloroplast DNA data all suggest (a) origin of the monophyletic Hawaiian silversword alliance from a California tarweed ancestor, (b) closer relationship of the Hawaiian species to Madia and Raillardiopsis than to Adenothamnus or Raillardella, (c) paraphyly of Raillardiopsis, a segregate of Raillardella, and (d) closer relationship of Raillardiopsis to Madia and the silversword alliance than to Raillardella. These findings indicate that the ITS region in plants should be further explored as a promising source of nuclear phylogenetic markers.