Phylogenetic analysis of Asian<Emphasis Type="Italic"> Symplocos</Emphasis> (Symplocaceae) based on nuclear and chloroplast DNA sequences

Thirty species and one variety of Symplocos (Symplocaceae), including all taxa distributed in Japan, were phylogenetically analyzed with DNA sequence data. The evolution of morphological characters is discussed on the basis of the phylogenetic relationships obtained. All species were Asian, except one, S. austromexicana, from Mexico. The nuclear ribosomal internal transcribed spacer (ITS) region and two intergenic spacer regions, between trnL and trnF, and between trnH and psbA of chloroplast DNA were used. The topologies of trees obtained from ITS and the chloroplast intergenic spacers are largely congruent. S. sonoharae, the representative of the subgenus Symplocos in Asia, was sister to all other species. The position of the American species, S. austromexicana, which also belongs to the subgenus Symplocos, was not well resolved. The phylogenetic tree based on combined sequence data largely supports the monophyletic origin of the infrageneric sections proposed earlier. However, the phylogenetic relationship between them is not well resolved, probably due to rapid diversification. The section Palura, a deciduous group, is well defined in the DNA analysis, suggesting its independent status in the genus Symplocos. In spite of their morphological divergence, the three endemic species of the Bonin Islands are monophyletic. The occurrence of curved seeds seems to be homoplastic, scattered over the phylogenetic tree without showing a particular infrageneric relationship.     

A phylogeny inferred from large ribosomal subunit (26S) rDNA sequences suggests that Cuscuta is a derived member of Convolvulaceae

Cuscuta is a parasitic angiosperm that has been considered alternatively either as a genus within Convolvulaceae or as a monogeneric family in its own right. Although typically placed in the Solanales,Cuscuta has also been positioned within the Polemoniales. Extreme reduction of morphological and anatomical characters, as well as chloroplast genome reductions and rearrangements, has made the phylogenetic placement ofCuscuta uncertain. Analysis of 26S rDNA sequences suggests thatCuscuta is a derived member of Convolvulaceae. Molecular results are discussed in relation to the morphological and anatomical characters of autotrophic members of Convolvulaceae.     

绣球属(Hydrangea L.)植物分子系统学及系统发育分析

该研究基于对绣球属(Hydrangea L.)的大尺度取样,选取国内外61种绣球属和近缘属植物,分别基于核基因片段(ITS)和叶绿体基因片段(rbcL,trnL-F,atpB)重建了绣球属及其近缘种属的系统发育关系。结果表明:(1)核基因与叶绿体基因树之间在树形上没有明显的冲突,进而基于核基因和叶绿体基因联合数据重建了绣球属及其近缘种属的系统发育关系。(2)基于联合数据构建的系统树确认了2个大分支,并得到了果实顶端截平与否这一形态学证据的强力支持;每个大分枝又分为4个类群,共确定了8个类群。部分类群也得到了广义宏观形态性状的支持,如第1类群得到了叶形、花粉以及种子形态的支持。因此,该系统发育关系的重建对于全面理解绣球属及其近缘种属的演化关系具有重要的启发。     

Phylogeny and biogeography of Chinese and Australasian Polystichum ferns as inferred from chloroplast trnL-F and rps4-trnS sequence data

Polystichum, one of the largest genera of ferns, occurs worldwide with the greatest diversity in southwest China and adjacent regions. Although there have been studies of Chinese Polystichum on its traditional classification, geographic distributions, and even a few on its molecular systematics, its relationships to other species outside China remain little known. Here, we investigated the phylogeny and biogeography of the Polystichum species from China and Australasia. The evolutionary relationships among 42 Polystichum species found in China (29 taxa) and Australasia (13 taxa) were inferred from phylogenetic analyses of two chloroplast DNA sequence data sets: rps4-trnS and trnL-F intergenic spacers. The divergence time between Chinese and Australasian Polystichum was estimated. The results indicated that the Australasian species comprise a monophyletic group that is nested within the Chinese diversity, and that the New Zealand species are likewise a monophyletic group nested within the Australasian species. The divergence time estimates suggested that Chinese Polystichum migrated into Australasia from around 40 Ma ago, and from there to New Zealand from about 14 Ma. The diversification of the New Zealand Polystichum species began about 10 Ma. These results indicated that Polystichum probably originated in eastern Asia and migrated into Australasia: first into Australia and then into New Zealand.     

Confirmation of natural hybrids between Gentiana straminea and G. siphonantha (Gentianaceae) based on molecular evidence

A few individuals with intermediate morphology always appeared in the sympatric distributions of Gentiana straminea and G. siphonantha. These intermediate individuals were hypothesized to be the hybrids of two species after a careful evaluation of their morphological characteristics. To test this hypothesis, sequence comparison of the internal transcribed spacer (ITS) regions of the nuclear ribosomal and trnS (GCU)-trnG (UCC) intergenic spacer region of the chloroplast DNA from Gentiana straminea, G. siphonantha and the putative hybrids was performed. The results suggest that most intermediate individuals were the natural hybrids between G. straminea and G. siphonantha. In addition, we examined the sequence variation among the individuals of both parent species and analyzed the possibility leading to the incongruent identification in some individuals based on morphologic and molecular evidences, respectively. The intraspecific diversification of DNA fragments within both parent species and their high variability in hybrid swarms probably resulted from chloroplast genome recombination and incomplete lineage sorting during the early stages of speciation origin of the parent species. __________ Translated from Acta Botanica Yunnanica, 2007, 29 (1): 91–97 [译自:云南植物研究]     

Unknown New Zealand angiosperm assigned toCunoniaceae using sequence of the chloroplastrbcL gene

We report the first example of identification of an unknown flowering plant to family level using comparison of DNA sequences. The newly-discovered New Zealand plant had not been identified by traditional methods, but has been placed in the Gondwanic familyCunoniaceae on the basis of the sequence of its chloroplastrbcL gene. It has not flowered, but its vegetative characters are typical of the family. However, it is clearly unusual in several morphological and anatomical features and appears to match no known genus. We consider that, although it probably represents an unnamed new genus, it could possibly be a mutant dwarf form ofWeinmannia racemosa, so it is not prudent to name it at this time.     

The phylogenetic utility of chloroplast and nuclear DNA markers and the phylogeny of the Rubiaceae tribe Spermacoceae

The phylogenetic utility of chloroplast (atpB-rbcL, petD, rps16, trnL-F) and nuclear (ETS, ITS) DNA regions was investigated for the tribe Spermacoceae of the coffee family (Rubiaceae). ITS was, despite often raised cautions of its utility at higher taxonomic levels, shown to provide the highest number of parsimony informative characters, in partitioned Bayesian analyses it yielded the fewest trees in the 95% credible set, it resolved the highest proportion of well resolved clades, and was the most accurate region as measured by the partition metric and the proportion of correctly resolved clades (well supported clades retrieved from a combined analysis regarded as “true”). For Hedyotis, the nuclear 5S-NTS was shown to be potentially as useful as ITS, despite its shorter sequence length. The chloroplast region being the most phylogenetically informative was the petD group II intron.We also present a phylogeny of Spermacoceae based on a Bayesian analysis of the four chloroplast regions, ITS, and ETS combined. Spermacoceae are shown to be monophyletic. Clades supported by high posterior probabilities are discussed, especially in respect to the current generic classification. Notably, Oldenlandia is polyphyletic, the two subgenera of Kohautia are not sister taxa, and Hedyotis should be treated in a narrow sense to include only Asian species.     

Homologues of the green algal gidA gene and the liverwort frxC gene are present on the chloroplast genomes of conifers

Strong hybridization signals were obtained from total DNA of two conifers, lodgepole pine (Pinus contorta) and Norway spruce (Picea abies), in a Southern blot analysis using a probe derived from the chloroplast gidA gene of the green alga Chlamydomonas reinhardtii. The pine fragments detected by the probe were found to originate from the chloroplast genome and, as judged by the signal intensity, this was also true for the spruce fragments. Sequence analysis of the hybridizing pine chloroplast DNA region revealed an open reading frame potentially encoding a 459 amino acid polypeptide, highly homologous to that deduced from the algal gene and to ORF465 of liverwort chloroplast DNA. Upstream of the gidA sequence, we found a trnN(GUU) gene and an open reading frame of 291 codons which was 78% identical to the frxC gene of liverwort. Since ORF465 is located immediately downstream of trnN and frxC in liverwort, the genetic organization of this region is very similar in the two plants. In contrast, neither the gidA nor the frxC gene is present in the chloroplast DNA of tobacco or rice. It was recently reported that deletions in the gidA region of the chloroplast genome of Chlamydomonas reinhardtii abolish the light-independent pathway of chlorophyll synthesis which exists in many algae and lower plants. The presence of the gidA gene on the chloroplast genomes of conifers may therefore be of significance with respect to the ability of these plants to synthesize chlorophyll in the dark.     

Chloroplast DNA of Acetabularia mediterranea: Cell cycle related changes in distribution

Chloroplast DNA (cpDNA) distribution in the giant unicellular, uninucleate alga Acetabularia mediterranea was analyzed with the DNA-specific fluorochrome 4'6-diamidino-2-phenylindole (DAPI) at various stages of the cell cycle. The number of chloroplasts exhibiting DNA/DAPI fluorescence changes during the cell's developmental cycle: (1) all chloroplasts in germlings contain DNA; (2) the number of plastids with DNA declines during polar growth of the vegetative cell; (3) it increases again prior to the transition from the vegetative to the generative phase; (4) several nucleoids of low fluorescence intensity are present in the chloroplasts of the gametes. The temporal distribution of the number of chloroplasts with DNA appears to be linked to the different mode of chloroplast division and growth during the various stages of development. The chloroplast cycle in relation to the cell cycle is discussed.Abbreviations cpDNA chloroplast DNA - DAPI 4,6-diamidino-2-phenylindole     

Nicotiana cybrids with Petunia chloroplasts

Summary Protoplasts of a chloroplast-defective cultivar of Nicotiana tabacum were fused with gamma-irradiated protoplasts of Petunia hybrida. Over 100 photoautotrophic plants were regenerated; of these 94 were tested for Petunia chloroplast traits and all but one had Petunia chloroplasts based on their sensitivity to the fungal toxin, tentoxin. Chloroplast DNA was analysed for 3 of the sensitive plants and was shown to be identical to Petunia chloroplast DNA. Most of the plants (about 70%) appeared to be normal N. tabacum plants, based on morphology and chromosome number. They were fully fertile with normal pollen viability, seed set, and seed viability. The remaining 30% of the plants showed varying degrees of vegetative and reproductive abnormalities.The techniques of somatic cell genetics have led to many possible nuclear-organellar combinations that may be considered as cybrids. In this paper, we use the term to include the combination of nucleus from one species and chloroplast from another species     

Chloroplast DNA insertions into the nuclear genome of rice: the genes,sites and ages of insertion involved

Rice (Oryza sativa) is one of three predominant grain crops, and its nuclear and organelle genomes have been sequenced. Following genome analysis revealed many exchanges of DNA sequences between the nuclear and organelle genomes. In this study, a total of 45 chloroplast DNA insertions more than 2 kb in length were detected in rice nuclear genome. A homologous recombination mechanism is expected for those chloroplast insertions with high similarity between their flanking sequences. Only five chloroplast insertions with high sequence similarity between two flanking sequences from an insertion were found in the 45 insertions, suggesting that rice might follow the non-homologous end-joining (NHEJ) repair of double-stranded breaks mechanism, which is suggested to be common to all eukaryotes. Our studies indicate that the most chloroplast insertions occurred at a nuclear region characterized by a sharp change of repetitive sequence density. One potential explanation is that regions such as this might be susceptible target sites or “hotspots” of DNA damage. Our results also suggest that the insertion of retrotransposon elements or non-chloroplast DNA into chloroplast DNA insertions may contribute significantly to their fragmentation process. Moreover, based on chloroplast insertions in nuclear genomes of two subspecies (indica and japonica) of cultivated rice, our results strongly suggest that they diverged during 0.06–0.22 million years ago. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Mapping of a chloroplast RFLP marker associated with the CMS cytoplasm of sugar beet (Beta vulgaris)

The Owen cytoplasm of male-sterile sugar beet is associated with several alterations of mitochondrial DNA and one additional HindIII site of chloroplast DNA. The region of this HindIII site has been cloned and sequenced. The site maps in a small reading frame (orf32) close to the ycf7 (orf31) gene in the petG-psbE region of chloroplast DNA. Possible functional implications of the results are discussed. The chloroplast RFLP marker described could be useful for studies on chloroplast-mitochondrial interactions, CMS of sugar beet, and the origin of the Owen cytoplasm.     

Molecular Phylogenetics and Biogeography of the Caribbean-Centered <Emphasis Type="Italic">Croton</Emphasis> Subgenus <Emphasis Type="Italic">Moacroton</Emphasis> (Euphorbiaceae s.s.)

Initial molecular phylogenetic studies established the monophylly of the large genus Croton (Euphorbiaceae s.s.) and suggested that the group originated in the New World. A denser and more targeted sampling of Croton species points to a South American origin for the genus. The nuclear and chloroplast genomes indicate a different rooting for the phylogeny of Croton. Although we favor the rooting indicated by the chloroplast data our conclusions are also consistent with the topology inferred from the nuclear data. The satellite genera Cubacroton and Moacroton are embedded within Croton. These two genera are synonimized into Croton and a new subgenus, Croton subgenus Moacroton, is circumscribed to include them and their allied Croton species. Croton subgenus Moacroton is morphologically characterized by a primarily lepidote indumentum, bifid or simple styles, and pistillate flowers with sepals that are connate at the base. This newly circumscribed subgenus is found from North America to South America, and in contrast to the majority of Croton species most of its members are found in mesic habitats. The group is most diverse in the greater Caribbean basin. A molecular clock was calibrated to the phylogeny using the available Euphorbiaceae fossils. The timing and pattern of diversification of Croton is consistent with both the GAARlandia and Laurasian migration hypotheses. A single species, Croton poecilanthus from Puerto Rico, is placed incongruently by its nuclear and chloroplast genomes. The possibility of this species being of hybrid origin is discussed.      

Chloroplast DNA of black pine retains a residual inverted repeat lacking rRNA genes: nucleotide sequences of trnQ, trnK, psbA, trnI and trnH and the absence of rps16

Summary A physical map of black pine (Pinus thunbergii) chloroplast DNA (120 kb) was constructed and two separate portions of its nucleotide sequence were determined. One portion contains trnQ-UUG, ORF510, ORF83, trnK-UUU (ORF515 in the trnK intron), ORF22, psbA, trnI-CAU (on the opposing strand) and trnH-GUG, in that order. Sequence analysis of another portion revealed the presence of a 495 by inverted repeat containing trnI-CAU and the 3 end of psbA but lacking rRNA genes. The position of trnI-CAU is unique because most chloroplast DNAs have no gene between psbA and trnH (trnI-CAU is usually located further downstream). Black pine chloroplast DNA lacks rps16, which has been found between trnQ and trnK in angiosperm chloroplast DNAs, but possesses ORF510 instead. This ORF is highly homologous to ORF513 found in the corresponding region of liverwort chloroplast DNA and ORF563 located downstream from trnT in Chlamydomonas moewusii chloroplast DNA. A possible pathway for the evolution of black pine chloroplast DNA is discussed.     

Comparison of chloroplast and nuclear phylogeny in the autogamous annualMicroseris douglasii (Asteraceae: Lactuceae)

Morphology suggests that the Californian annualMicroseris douglasii is a monophyletic sister group to the other three diploid annuals ofMicroseris. Phylogenetic analysis of 44 inbred strains ofM. douglasii derived from 23 populations with 72 RAPD markers in the nuclear DNA strongly supports this phylogeny. However, 13 chloroplast RFLPs divideM. douglasii into four distinct groups. Two of these each share one or more cpRFLPs withM. bigelovii andM. pygmaea. Several hypotheses can explain the incongruence between nuclear and chloroplast phylogeny: (1) random sorting out of chloroplasts during phylogeny from a polymorphic pool, (2) cytoplasmic introgression from the related annualM. bigelovii intoM. douglasii after hybridization followed by elimination of theM. bigelovii nuclear genome. We suggest cytoplasmic introgression as the most likely origin. Possible remnants of nuclear introgression have been found in two populations ofM. douglasii that are polymorphic for chloroplast types. In these populationsM. bigelovii type chloroplast DNA seems to be accompanied by nuclear genes for flower color and leaf shape.     

Hybridisation processes in sympatric populations of pines Pinus sylvestris L., P. mugo Turra and P. uliginosa Neumann

Natural hybridisation was postulated between the closely related pine species Pinus sylvestris and the P. mugo complex, however no clear evidence on propagation of mature hybrids in nature has been documented so far. To test the hybridisation hypothesis we applied chloroplast DNA (cpDNA) markers and isozymes in the analyses of 300 individuals representing the variety of morphological forms in the sympatric populations of P. sylvestris, P. mugo and P. uliginosa at the peat bog complex in the Sudety Mts., Poland. Additionally, the haplotypes of paternally inherited cpDNA of 149 open pollinated progeny derived from seeds were compared to the haplotypes of parental trees to access the intensity and direction of contemporary hybridisation. The morphologically highly variable polycormic (multi-stemmed) hybrids between P. mugo and P. uliginosa were identified. The second group of hybrids was found among the monocormic (single-stemmed) P. sylvestris-like individuals carrying the cpDNA from P. mugo complex. Hybrids of P. sylvestris as a pollen donor and P. mugo or P. uliginosa as a mother were not found, either in the group of examined trees, or among the open pollinated progeny. The results indicate that numerous hybrids can exist in the sympatric population of the species studied and that gene flow can successfully proceed from P. mugo complex to P. sylvestris. Hybridisation and ecological selection seems to play a significant role in diversification and evolution of the investigated species.     

Molecular and genetic analysis of the chloroplast ATPase of chlamydomonas

Summary We have carried out a molecular and genetic analysis of the chloroplast ATPase in Chlamydomonas reinhardtii. Recombination and complementation studies on 16 independently isolated chloroplast mutations affecting this complex demonstrated that they represent alleles in five distinct chloroplast genes. One of these five, the ac-u-c locus, has been positioned on the physical map of the chloroplast DNA by deletion mutations. The use of cloned spinach chloroplast ATPase genes in heterologous hybridizations to Chlamydomonas chloroplast DNA has allowed us to localize three or possibly four of the ATPase genes on the physical map. The beta and probably the epsilon subunit genes of Chlamydomonas CF₁ lie within the same region of chloroplast DNA as the ac-u-c locus, while the alpha and proteolipid subunit genes appear to map adjacent to one another approximately 20 kbp away. Unlike the arrangement in higher plants, these two pairs of genes are separated from each other by an inverted repeat.     

Chloroplast DNA variation and population structure in the widespread forest tree, Eucalyptus grandis

Recognition of genetic structure of populations and the ability to identify vulnerable populations is useful for the formation of conservation management strategies for plants. Eucalyptus grandis is a tall forest tree that has a major area of occurrence in subtropical eastern Australia, with smaller populations located in the east coast tropics. Many widespread forest species exhibit population differentiation that corresponds to geographic regions. However, Eucalyptus grandis appears to be an exception based on isozyme and morphological data. This is intriguing given a large discontinuity between northern populations and those in the southern part of the species range. In this study, the distribution of a maternally inherited chloroplast locus was examined because it was more likely to reveal genetic structure due to the slower evolution of the chloroplast genome and limited dispersal of seed in eucalypts. As expected, the G _ST for chloroplast DNA was higher than that for nuclear DNA but indicated low population differentiation for a forest tree species. Phylogeographic analysis indicated that the 15 populations grouped into three broad geographical regions; however, overall population structure was weak suggesting that the large geographical disjunction in the distribution of E. grandis may be relatively recent. A paradigm for conservation management of E. grandis based on chloroplast DNA haplotype distribution would take into account the low differentiation among populations.     

Chloroplast DNA restriction site mapping and the phylogeny ofRanunculus (Ranunculaceae)

A chloroplast DNA restriction site map forRanunculus sceleratus (Ranunculaceae) was constructed using 14 restriction endonucleases. The total size of the chloroplast genome is 152.4kb. No inversions were detected relative to the tobacco chloroplast DNA. Cladistic analyses of chloroplast DNA restriction site polymorphism were employed in order to elucidate the phylogeny among 76 species of the genusRanunculus in a wide sense and one species ofTrautvetteria. A total of 341 informative restriction site changes were detected. Parsimony jackknifing, bootstrapping and decay analysis were undertaken in order to evaluate the amount of support for the monophyletic groups. The results suggest that the analysed species ofRanunculus are divisible into two main clades. Only few of the traditional sections and subgenera ofRanunculus are monophyletic. The genusTrautvetteria is nested within a clade comprising, e.g.Ranunculus cymbalaria, R. andersonii, R. lapponicus andR. ficaria. SubgenusBatrachium lies within a larger clade containing, e.g.R. sceleratus andR. hyperboreus. Contractions of the inverted repeat due to parallel deletions of 200–300 bp close to the J_SB have occurred in many clades and the phylogenetic distribution of this size reduction was mapped among the species.