Phylogenetics of Pinus (Pinaceae) based on nuclear ribosomal DNA internal transcribed spacer region sequences

A 650-bp portion of the nuclear ribosomal DNA internal transcribed spacer region was sequenced in 47 species of Pinus, representing all recognized subsections of the genus, and 2 species of Picea and Cathaya as outgroups. Parsimony analyses of these length variable sequences were conducted using a manual alignment, 13 different automated alignments, elision of the automated alignments, and exclusion of all alignment ambiguous sites. High and moderately supported clades were consistently resolved across the different analyses, while poorly supported clades were inconsistently recovered. Comparison of the topologies highlights taxa of particularly problematic placement including Pinus nelsonii and P. aristata. Within subgenus Pinus, there is moderate support for the monophyly of a narrowly circumscribed subsect. Pinus (=subsect. Sylvestres) and strong support for a clade of North and Central American hard pines. The Himalayan P. roxburghii may be sister species to these "New World hard pines," which have two well-supported subgroups, subsect. Ponderosae and a clade of the remaining five subsections. The position of subsect. Contortae conflicts with its placement in a chloroplast DNA restriction site study. Within subgenus Strobus there is consistent support for the monophyly of a broadly circumscribed subsect. Strobi (including P. krempfii and a polyphyletic subsect. Cembrae) derived from a paraphyletic grade of the remaining soft pines. Relationships among subsects. Gerardianae, Cembroides, and Balfourianae are poorly resolved. Support for the monophyly of subgenus Pinus and subgenus Strobus is not consistently obtained.     

基于matK基因的松属白皮松组分子系统发育分析

基于matK基因对松属(Pinus L.)白皮松组(sect.Parrya Myre)进行了分子系统发育分析.白皮松组为一个并系类群,因为白松组的成员与该组(包括越南的扁叶松(P.krempfii Lecomte))的亚洲成员形成一个强烈支持的分支(靴带值92%).在这个分支中,白松组的3个代表种形成一个稳定的单系,而白皮松组的亚洲成员之间系统发育关系不明确.扁叶松和西藏白皮松(P. gerardiana Wall.ex D.Don)聚在一起,但只有61%的支持率.虽然在以前4个cpDNA基因序列分析时五针白皮松(P.squamata X.W.Li)与白皮松(P.bungeana Zucc.ex Loud.)和西藏白皮松形成一个单系,但在本文的分析中三者的关系不明确.在邻接树和多数一致简约树上,北美的白皮松组成员形成一个支持率低的分支.北美的subsect.Balfourianae Engelm.亚组(包括P.aristata Engelm.)是一个单系,但支持率较低.美洲另外两个亚组subsect.Cembroides Englem.和subsect.Rzedowskianae Carv.的组间和组内关系不确定,它们在严格一致简约树上形成一个多歧分支.     

基于matK基因的松属白皮松组分子系统发育分析(英文)

基于matK基因对松属(Pinus L.)白皮松组(sect.Parrya Myre)进行了分子系统发育分析。白皮松组为一个并系类群,因为白松组的成员与该组(包括越南的扁叶松(P.krempfii Lecomte))的亚洲成员形成一个强烈支持的分支(靴带值92％)。在这个分支中,白松组的3个代表种形成一个稳定的单系,而白皮松组的亚洲成员之间系统发育关系不明确。扁叶松和西藏白皮松(P.gerardiana Wall.ex D.Don)聚在一起,但只有61％的支持率。虽然在以前4个cpDNA基因序列分析时五针白皮松(P.squamata X.W.Li)与白皮松(P.bungeana Zucc.ex Loud.)和西藏白皮松形成一个单系,但在本文的分析中三者的关系不明确。在邻接树和多数一致简约树上,北美的白皮松组成员形成一个支持率低的分支。北美的subsect.Balfourianae Engelm.亚组(包括P.aristata Engelm.)是一个单系,但支持率较低。美洲另外两个亚组subsect.Cembroides Englem.和subsect.Rzedowskianae Carv.的组间和组内关系不确定,它们在严格一致简约树上形成一个多歧分支。     

Complete cpDNA genome sequence of Smilax china and phylogenetic placement of Liliales--influences of gene partitions and taxon sampling

The complete nucleotide sequence of the chloroplast genome (cpDNA) of Smilax china L. (Smilacaceae) is reported. It is the first complete cp genome sequence in Liliales. Genomic analyses were conducted to examine the rate and pattern of cpDNA genome evolution in Smilax relative to other major lineages of monocots. The cpDNA genomic sequences were combined with those available for Lilium to evaluate the phylogenetic position of Liliales and to investigate the influence of taxon sampling, gene sampling, gene function, natural selection, and substitution rate on phylogenetic inference in monocots. Phylogenetic analyses using sequence data of gene groups partitioned according to gene function, selection force, and total substitution rate demonstrated evident impacts of these factors on phylogenetic inference of monocots and the placement of Liliales, suggesting potential evolutionary convergence or adaptation of some cpDNA genes in monocots. Our study also demonstrated that reduced taxon sampling reduced the bootstrap support for the placement of Liliales in the cpDNA phylogenomic analysis. Analyses of sequences of 77 protein genes with some missing data and sequences of 81 genes (all protein genes plus the rRNA genes) support a sister relationship of Liliales to the commelinids-Asparagales clade, consistent with the APG III system. Analyses of 63 cpDNA protein genes for 32 taxa with few missing data, however, support a sister relationship of Liliales (represented by Smilax and Lilium) to Dioscoreales-Pandanales. Topology tests indicated that these two alignments do not significantly differ given any of these three cpDNA genomic sequence data sets. Furthermore, we found no saturation effect of the data, suggesting that the cpDNA genomic sequence data used in the study are appropriate for monocot phylogenetic study and long-branch attraction is unlikely to be the cause to explain the result of two well-supported, conflict placements of Liliales. Further analyses using sufficient nuclear data remain necessary to evaluate these two phylogenetic hypotheses regarding the position of Liliales and to address the causes of signal conflict among genes and partitions.     

Molecular Phylogeny of Section Parrya of Pinus （Pinaceae） Based on Chloroplast matK Gene Sequence Data

The molecular phylogenetics of sect. Parrya Myre of Pinus L. was analyzed based onchloroplast matKgene sequence data. The section was resolved as paraphyletic because members of thesect. Strobus were nested within a clade composed by the Asian members of the section, including theVietnamese P. krempfii Lecomte, which was strongly supported with a bootstrap value of 92%. [n thistopology, the three sampled species of sect. Strobus formed a strongly supported monophyletic group,while their relationships of Asian species of sect. Parrya were not clear. P. krempfii was grouped with P.gerardiana Wall. ex D. Don with low bootstrap support. The relationships among the Asian members of thesect. Parrya, i.e.P, bungeana Zucc. ex Loud., P. gerardiana and the recently described endangered pine, P.squarnata X. W. Li, was not resolved, although the monophyly of the three pines was strongly supported inthe combined analysis of four cpDNA sequences. The topology of the neighbor joining tree revealed anassemblage of the American members of the section, which also appeared in the majority rule tree withweak bootstrap support. However, this assemblage was not resolved in the consensus tree of theparsimonious analysis. The American subsect. Ba/fourianae Engelm. formed a weakly supported groupincluding P. aristata Engelm., while the relationships among and within the other two American subsections,Cembroides Englem. and tTzedowskianae Carv., were not resolved, as the members of them formed apolytomy in the consensus tree of the parsimonious analysis. The biogeographical implications of theresults are also discussed in this paper.     

Widespread genealogical nonmonophyly in species of Pinus subgenus Strobus

Phylogenetic relationships among Pinus species from subgenus Strobus remain unresolved despite combined efforts based on nrITS and cpDNA. To provide greater resolution among these taxa, a 900-bp intron from a late embryogenesis abundant (LEA)-like gene (IFG8612)was sequenced from 39 pine species, with two or more alleles representing 33 species. Nineteen of 33 species exhibited allelic nonmonphyly in the strict consensus tree, and 10 deviated significantly from allelic monophyly based on topology incongruence tests. Intraspecific nucleotide diversity ranged from 0.0 to 0.0211, and analysis of variance shows that nucleotide diversity was strongly associated (P < 0.0001)with the degree of species monophyly. Although species nonmonophyly complicates phylogenetic interpretations, this nuclear locus offers greater topological support than previously observed for cpDNA or nrITS. Lacking evidence for hybridization, recombination, or imperfect taxonomy, we feel that incomplete lineage sorting remains the best explanation for the polymorphisms shared among species. Depending on the species, coalescent expectations indicate that reciprocal monophyly will be more likely than paraphyly in 1.71 to 24.0 x 10(6) years, and that complete genome-wide coalescence in these species may require up to 76.3 x 10(6) years. The absence of allelic coalescence is a severe constraint in the application of phylogenetic methods in Pinus, and taxa sharing similar life history traits with Pinus are likely to show species nonmonophyly using nuclear markers.     

Paralogy and orthology in the MALVACEAE rpb2 gene family: investigation of gene duplication in hibiscus

A sample of the second largest subunit of low-copy nuclear RNA polymerase II (rpb2) sequences from Malvaceae subfamily Malvoideae suggests that rpb2 has been duplicated early in the subfamily's history. Hibiscus and related taxa possess two rpb2 genes, both of which produce congruent phylogenetic patterns that are largely concordant with cpDNA topologies. No evidence of functional divergence or disruption was found among duplicated copies, suggesting that long-term maintenance of duplicated copies of rpb2 is usual in this lineage. Therefore, this gene may be suitable for the potential diagnosis of relatively old polyploid events. One probable pseudogene was found in Radyera farragei and a single chimeric sequence was recovered from Howittia trilocularis, suggesting that the rpb2 locus is not as prone to evolutionary processes that can confound phylogenetic inferences based on nDNA sequences. The pattern of relationships among rpb2 sequences, coupled with chromosome number information and Southern hybridization data, suggests that an early polyploid event was not the cause of the duplication, despite independent evidence of paleopolyploidy in some members of Malvoideae. Rpb2 exons and introns together are suitable for phylogenetic analysis, producing well-resolved and well-supported results that were robust to model permutation and congruent with previous studies of subfamily Malvoideae using cpDNA characters.     

Molecular phylogeny of Cotoneaster (Rosaceae) inferred from nuclear ITS and multiple chloroplast sequences

Cotoneaster Medik. (Rosaceae, Maloideae) is distributed in Europe, North Africa, and temperate areas of Asia except Japan. Members of the genus exhibit considerable morphological variation. The infrageneric classification is also obscured by polyploidy, hybridization, and apomixis. In this study, phylogenetic analyses were conducted to test infrageneric classifications of this genus using DNA sequence data from the nuclear ITS (nrITS) region and three chloroplast intergenic spacer regions. Maximum parsimony and Bayesian inference analyses of both datasets agreed with the two sections/subgenera of Koehne’s classification system, and suggested that four subsections (Microphylli, Chaenopetalum, Adpressi, and Cotoneaster) and the series of Koehne’s classification system were non-monophyletic. The incongruence length difference test indicated that the nrITS and cpDNA datasets were significantly incongruent (P = 0.001), and the placement of 14 species was discordant in phylogenetic trees derived from the two datasets. Within Cotoneaster, hybridization was indicated to be an important factor contributing to the incongruence between the nrITS and cpDNA data. By mapping nine morphological characters onto the combined nrITS–cpDNA phylogenetic tree, we inferred that a deciduous habit, glabrous fruit, white anthers, erect and light pink petals, and white filaments are plesiomorphic character states in Cotoneaster.     

Large-scale asymmetric introgression of cytoplasmic DNA reveals Holocene range displacement in a North American boreal pine complex

Jack pine (Pinus banksiana) and lodgepole pine (Pinus contorta var. latifolia) are two North American boreal hard pines that hybridize in their zone of contact in western Canada. The main objective of this study was to characterize their patterns of introgression resulting from past and recent gene flow, using cytoplasmic markers having maternal or paternal inheritance. Mitochondrial DNA (mtDNA) and chloroplast DNA (cpDNA) diversity was assessed in allopatric populations of each species and in stands from the current zone of contact containing morphological hybrids. Cluster analyses were used to identify genetic discontinuities among groups of populations. A canonical analysis was also conducted to detect putative associations among cytoplasmic DNA variation, tree morphology, and site ecological features. MtDNA introgression was extensive and asymmetric: it was detected in P. banksiana populations from the hybrid zone and from allopatric areas, but not in P. contorta populations. Very weak cpDNA introgression was observed, and only in P. banksiana populations. The mtDNA introgression pattern indicated that central Canada was first colonized by migrants from a P. contorta glacial population located west of the Rocky Mountains, before being replaced by P. banksiana migrating westward during the Holocene. In contrast, extensive pollen gene flow would have erased the cpDNA traces of this ancient presence of P. contorta. Additional evidence for this process was provided by the results of canonical analysis, which indicated that the current cpDNA background of trees reflected recent pollen gene flow from the surrounding dominant species rather than historical events that took place during the postglacial colonization.     

Utility of Low-Copy Nuclear Gene Sequences in Plant Phylogenetics

Low-copy nuclear genes in plants are a rich source of phylogenetic information. They hold a great potential to improve the robustness of phylogenetic reconstruction at all taxonomic levels, especially where universal markers such as cpDNA and nrDNA are unable to generate strong phylogenetic hypotheses. Low-copy nuclear genes, however, remain underused in plant phylogenetic studies due to practical and theoretical complications in unraveling the evolutionary dynamics of nuclear gene families. The lack of the universal markers or universal PCR primers of low-copy nuclear genes has also hampered their phylogenetic utility. It has recently become clear that low-copy nuclear genes are particularly helpful in resolving close interspecific relationships and in reconstructing allopolyploidization in plants. Gene markers that are widely, if not universally, useful have begun to emerge. Although utilizing low-copy nuclear genes usually requires extra lab work such as designing PCR primers, PCR-cloning, and/or Southern blotting, rapid accumulation of gene sequences in the databases and advances in cloning techniques have continued to make such studies more feasible. With the growing number of theoretical studies devoted to the gene tree and species tree problem, a solid foundation for reconstructing complex plant phylogenies based on multiple gene trees began to build. It is also realized increasingly that fast evolving introns of the low-copy nuclear genes will provide much needed phylogenetic information around the species boundary and allow us to address fundamental questions concerning processes of plant speciation. Phylogenetic and molecular evolutionary analyses of developmentally important genes will add a new dimension to systematic and evolutionary studies of plant diversity.     

Variation in the nrDNA ITS of Pinus subsection Cembroides: implications for molecular systematic studies of pine species complexes.

The pinyon pines (Pinus subsection Cembroides), distributed in semiarid regions of the western United States and Mexico, include a mixture of relictual and more recently evolved taxa. To investigate relationships among the pinyons, we screened and partially sequenced 3000-bp clones of the nuclear ribosomal DNA internal transcribed spacer (ITS) region for 16 taxa from subsect. Cembroides and nine representatives from four other subsections of subgenus Strobus. Restriction digests of clones reveal within-individual heterogeneity, suggesting that concerted evolution is operating slowly on the ITS in pine species. Two ITS clones were identified as pseudogenes. Tandem subrepeats in the ITS1 form stem loops comparable to those in other genera of Pinaceae and may be promoting recombination between rDNA repeats, resulting in ITS1 chimeras. Within the pinyon clade, phylogenetic structure is present, but different clones from the same (or different) individuals of a species are polyphyletic, indicating that coalescence of ITS copies within individual genomes predates evolutionary divergence in the group. At the level of subsection and above, the ITS region corresponds well with morphological and cpDNA evidence. Except for P. nelsonii, the pinyons are monophyletic, with both subsect. Cembroides and P. nelsonii forming a clade with the foxtail and bristlecone pines (subsect. Balfourianae) of western North America.     

Utility of low-copy nuclear gene sequences in plant phylogenetics

Low-copy nuclear genes in plants are a rich source of phylogenetic information. They hold a great potential to improve the robustness of phylogenetic reconstruction at all taxonomic levels, especially where universal markers such as cpDNA and nrDNA are unable to generate strong phylogenetic hypotheses. Low-copy nuclear genes, however, remain underused in plant phylogenetic studies due to practical and theoretical complications in unraveling the evolutionary dynamics of nuclear gene families. The lack of the universal markers or universal PCR primers of low-copy nuclear genes has also hampered their phylogenetic utility. It has recently become clear that low-copy nuclear genes are particularly helpful in resolving close interspecific relationships and in reconstructing allopolyploidization in plants. Gene markers that are widely, if not universally, useful have begun to emerge. Although utilizing low-copy nuclear genes usually requires extra lab work such as designing PCR primers, PCR-cloning, and/or Southern blotting, rapid accumulation of gene sequences in the databases and advances in cloning techniques have continued to make such studies more feasible. With the growing number of theoretical studies devoted to the gene tree and species tree problem, a solid foundation for reconstructing complex plant phylogenies based on multiple gene trees began to build. It is also realized increasingly that fast evolving introns of the low-copy nuclear genes will provide much needed phylogenetic information around the species boundary and allow us to address fundamental questions concerning processes of plant speciation. Phylogenetic and molecular evolutionary analyses of developmentally important genes will add a new dimension to systematic and evolutionary studies of plant diversity.     

Incongruence among mitochondrial,chloroplast and nuclear gene trees in <Emphasis Type="Italic">Pinus</Emphasis> subgenus <Emphasis Type="Italic">Strobus</Emphasis> (Pinaceae)

Introgression has been considered to be one of main factors leading to phylogenetic incongruence among different datasets at lower taxonomic levels. In the plants of Pinaceae, the mtDNA, cpDNA, and nuclear DNA (nrDNA) may have different evolutionary histories through introgression because they are inherited maternally, paternally and biparentally, respectively. We compared mtDNA, cpDNA, and two low-copy nrDNA phylogenetic trees in the genus Pinus subgenus Strobus, in order to detect unknown past introgression events in this group. nrDNA trees were mostly congruent with the cpDNA tree, and supported the recent sectional and subsectional classification system. In contrast, mtDNA trees split the members of sect. Quinquefoliae into two groups that were not observed in the other gene trees. The factors constituting incongruence may be divided into the following two categories: the different splits within subsect. Strobus, and the non-monophyly of subsect. Gerardianae. The former was hypothesized to have been caused by the past introgression of cpDNA, mtDNA or both between Eurasian and North American species through Beringia. The latter was likely caused by the chimeric structure of the mtDNA sequence of P. bungeana, which might have originated through past hybridization, or through a horizontal transfer event and subsequent recombination. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Five nuclear loci resolve the polyploid history of switchgrass (Panicum virgatum L.) and relatives

Polyploidy poses challenges for phylogenetic reconstruction because of the need to identify and distinguish between homoeologous loci. This can be addressed by use of low copy nuclear markers. Panicum s.s. is a genus of about 100 species in the grass tribe Paniceae, subfamily Panicoideae, and is divided into five sections. Many of the species are known to be polyploids. The most well-known of the Panicum polyploids are switchgrass (Panicum virgatum) and common or Proso millet (P. miliaceum). Switchgrass is in section Virgata, along with P. tricholaenoides, P. amarum, and P. amarulum, whereas P. miliaceum is in sect. Panicum. We have generated sequence data from five low copy nuclear loci and two chloroplast loci and have clarified the origin of P. virgatum. We find that all members of sects. Virgata and Urvilleana are the result of diversification after a single allopolyploidy event. The closest diploid relatives of switchgrass are in sect. Rudgeana, native to Central and South America. Within sections Virgata and Urvilleana, P. tricholaenoides is sister to the remaining species. Panicum racemosum and P. urvilleanum form a clade, which may be sister to P. chloroleucum. Panicum amarum, P. amarulum, and the lowland and upland ecotypes of P. virgatum together form a clade, within which relationships are complex. Hexaploid and octoploid plants are likely allopolyploids, with P. amarum and P. amarulum sharing genomes with P. virgatum. Octoploid P. virgatum plants are formed via hybridization between disparate tetraploids. We show that polyploidy precedes diversification in a complex set of polyploids; our data thus suggest that polyploidy could provide the raw material for diversification. In addition, we show two rounds of allopolyploidization in the ancestry of switchgrass, and identify additional species that may be part of its broader gene pool. This may be relevant for development of the crop for biofuels.     

Phylogenetic relationships of Eurasian pines (Pinus, Pinaceae) based on chloroplast rbcL, MATK, RPL20-RPS18 spacer, and TRNV intron sequences

The sequence divergence of chloroplast rbcL, matK, trnV intron, and rpl20-rps18 spacer regions was analyzed among 32 Pinus species and representatives of six other genera in Pinaceae. The total aligned sequence length is 3570 bp. Of the four sequences examined, matK evolved much faster than rbcL in Pinus and in other Pinaceae genera. The two noncoding regions did not show more divergence than the two coding regions, especially within each Pinus subgenus. Phylogenetic analyses based on these four sequences gave consistent results and strongly supported the monophyly hypothesis for the genus Pinus and its two recognized subgenera. Pinus krempfii, the two-flat-needle pine endemic to Vietnam, was placed in subgen. Strobus and showed closer affinity to subsect. Gerardianae. The ancient character of sect. Parrya is further confirmed. However, monophyly of the sect. Parrya is not supported by our data. Among the Eurasian pines of subgen. Pinus, Mediterranean pines formed one clade and the Asian members of subsect. Sylvestres formed another. The Himalayan P. roxburghii showed considerable divergence from all the other hard pines from both regions. Pinus merkusii was distinctly separated from all the Asian members of subsect. Sylvestres. The implications of our results for Pinus classification are discussed.     

Phylogeny of Campanuloideae (Campanulaceae) with Emphasis on the Utility of Nuclear Pentatricopeptide Repeat (PPR) Genes

Background

The Campanuloideae (Campanulaceae) are a highly diverse clade of angiosperms found mostly in the Northern Hemisphere, with the highest diversity in temperate areas of the Old World. Chloroplast markers have greatly improved our understanding of this clade but many relationships remain unclear primarily due to low levels of molecular evolution and recent and rapid divergence. Furthermore, focusing solely on maternally inherited markers such as those from the chloroplast genome may obscure processes such as hybridization. In this study we explore the phylogenetic utility of two low-copy nuclear loci from the pentatricopeptide repeat gene family (PPR). Rapidly evolving nuclear loci may provide increased phylogenetic resolution in clades containing recently diverged or closely related taxa. We present results based on both chloroplast and low-copy nuclear loci and discuss the utility of such markers to resolve evolutionary relationships and infer hybridization events within the Campanuloideae clade.

Results

The inclusion of low-copy nuclear genes into the analyses provides increased phylogenetic resolution in two species-rich clades containing recently diverged taxa. We also obtain support for the placement of two early diverging lineages (Jasione and Musschia-Gadellia clades) that have previously been unresolved. Furthermore, phylogenetic analyses of PPR loci revealed potential hybridization events for a number of taxa (e.g., Campanula pelviformis and Legousia species). These loci offer greater overall topological support than obtained with plastid DNA alone.

Conclusion

This study represents the first inclusion of low-copy nuclear genes for phylogenetic reconstruction in Campanuloideae. The two PPR loci were easy to sequence, required no cloning, and the sequence alignments were straightforward across the entire Campanuloideae clade. Although potentially complicated by incomplete lineage sorting, these markers proved useful for understanding the processes of reticulate evolution and resolving relationships at a wide range of phylogenetic levels. Our results stress the importance of including multiple, independent loci in phylogenetic analyses.     

Multi-gene analysis provides a well-supported phylogeny of Rosales

Despite many attempts to resolve evolutionary relationships among the major clades of Rosales, some nodes have been extremely problematic and have remained unresolved. In this study, we use two nuclear and 10 plastid loci to infer phylogenetic relationships among all nine families of Rosales. Rosales were strongly supported as monophyletic; within Rosales all family relationships are well-supported with Rosaceae sister to all other members of the order. Remaining Rosales can be divided into two subclades: (1) Ulmaceae are sister to Cannabaceae plus (Urticaceae+Moraceae); (2) Rhamnaceae are sister to Elaeagnaceae plus (Barbeyaceae+Dirachmaceae). One noteworthy result is that we recover the first strong support for a sister relationship between the enigmatic Dirachmaceae and Barbeyaceae. These two small families have distinct morphologies and potential synapomorphies remain unclear. Future studies should try to identify nonDNA synapomorphies uniting Barbeyaceae with Dirachmaceae.     

Deep divergences in the coffee family and the systematic position of Acranthera

Despite extensive efforts, there are unresolved questions on evolutionary relationships in the angiosperm family Rubiaceae. Here, information from six loci and 149 Rubiaceae taxa provide new insights. Acranthera and Coptosapelta are strongly supported as sisters. Pollen grains of Acranthera possess several features common in Rubiaceae, but amongst potential similarities with the unusual grains of Coptosapelta are the nature of the apertures and the structure of the sexine. Luculia, Acranthera and Coptosapelta are excluded from the three subfamilies Ixoroideae, Cinchonoideae and Rubioideae. Sipaneeae and Condamineeae form a clade, sister to remaining Ixoroideae. Rondeletieae and Guettardeae are sisters to remaining Cinchonoideae. Colletoecema is sister to remaining Rubioideae, followed by the Urophylleae–Ophiorrhizeae clade. Nuclear ITS provided structured information at all phylogenetic levels, but the main gain from adding nrITS was the increased resolution. Average support values also increased but were generally high also without nrITS and the increase was not statistically significant.     

Reticulate evolution in Thuja inferred from multiple gene sequences: implications for the study of biogeographical disjunction between eastern Asia and North America

The eastern Asia-North America disjunction is one of the most interesting biogeographical patterns, but its formation is still in much debate. Here nucleotide sequences of five cpDNA regions, nrDNA ITS and two low-copy nuclear genes (LEAFY, 4CL) were employed to reconstruct the phylogeny and to explore the historical biogeography of Thuja, a typical eastern Asia-North America disjunct genus. High topological discordance was observed between chloroplast and nuclear gene trees, even between different nuclear gene trees, suggesting that Thuja could have a reticulate evolutionary history due to multiple interspecific hybridization events. The eastern Asian species Thuja koraiensis might have obtained its chloroplast genome from the eastern North American species T. occidentalis by chloroplast capture, while the western North American species T. plicata is very likely to have inherited a recombinant cpDNA. Based on the phylogenetic analysis of multiple genes, DIVA-reconstruction of the distribution history, molecular clock estimation and fossil data, we inferred that Thuja could have originated from the high-latitude areas of North America in the Paleocene or earlier with subsequent expansion into eastern Asia through the Bering Land Bridge. The two eastern Asia species T. standishii and T. sutchuenensis have a sister relationship, and their split could have occurred in the Oligocene or early Miocene. In the present study, the selection of molecular markers in biogeographic studies was also discussed. Since most previous studies on the eastern Asia and North America disjunction are based on uniparentally inherited cpDNA and (or) directly sequenced nrDNA ITS data, the historical reticulate evolution in the studied groups might have been underestimated. Therefore, we suggest that multiple genes from different genomes, especially low-copy nuclear genes, be used in this research area in the future.     

Cryptic repeated genomic recombination during speciation in Gossypium gossypioides

The Mexican cotton Gossypium gossypioides is a perplexing entity, with conflicting morphological, cytogenetic, and molecular evidence of its phylogenetic affinity to other American cottons. We reevaluated the evolutionary history of this enigmatic species using 16.4 kb of DNA sequence. Phylogenetic analyses show that chloroplast DNA (7.3 kb), nuclear ribosomal internal transcribed spacers (ITS; 0.69 kb), and unique nuclear genes (8.4 kb) yield conflicting resolutions for G. gossypioides. Eight low-copy nuclear genes provide a nearly unanimous resolution of G. gossypioides as the basalmost American diploid cotton, whereas cpDNA sequences resolve G. gossypioides deeply nested within the American diploid clade sister to Peruvian G. raimondii, and ITS places G. gossypioides in an African (rather than an American) clade. These data, in conjunction with previous evidence from the repetitive fraction of the genome, implicate a complex history for G. gossypioides possibly involving temporally separated introgression events from genetically divergent cottons that are presently restricted to different hemispheres. Based on repetitive nuclear DNA, it appears that G. gossypioides experienced nuclear introgression from an African species shortly after divergence from the remainder of the American assemblage. More recently, hybridization with a Mexican species may have resulted in cpDNA introgression, and possibly a second round of cryptic nuclear introgression. Gossypium gossypioides provides a striking example of the previously unsuspected chimeric nature of some plant genomes and the resulting phylogenetic complexity produced by multiple historical reticulation events.