Nikolai Friesen

Molecular evidence for bicontinental hybridogenous genomic constitution in Lepidium sensu stricto (Brassicaceae) species from Australia and New Zealand.

Lepidium sensu stricto (s.s.) (Brassicaceae) (ca. 150 species) is distributed worldwide with endemic species on every continent. It is represented in Australia and New Zealand by 19 and seven native species, respectively. In the present study we used a nuclear ribosomal internal transcribed spacer (ITS) phylogeny in comparison with a cpDNA phylogeny to unravel the origin of Australian/New Zealand species. Although phylogenetic relationships within Lepidium s.s. were not fully resolved, the cpDNA data were in agreement with a Californian origin of Lepidium species from Australia/New Zealand. Strongly conflicting signals between the cp- and nuclear DNA phylogenetic analysis clearly indicated hybridogenous genomic constitution of Australian Lepidium s.s. species: All 18 studied Australian/New Zealand Lepidium s.s. species examined shared a Californian cpDNA type. While eleven Australian/New Zealand species appeared to harbor a Californian ITS type, a group of seven species shared a South African ITS type. This pattern is most likely explained by two trans-oceanic dispersals of Lepidium from California and Africa to Australia/New Zealand and subsequent hybridization followed by homogenization of the ribosomal DNA either to the Californian or South African ITS type in the two different lineages. Calibration of our molecular trees indicates a Pliocene/Pleistocene origin of Lepidium in Australia/New Zealand. Low levels of cpDNA and ITS sequence divergence and unresolved topologies within Australian/New Zealand species suggest a rapid and recent radiation of Lepidium after the hybridization event. This coincides with dramatic climatic changes in that geological epoch shaping the composition of the vegetation.

Contribution to ITS phylogeny of the Brassicaceae, with special reference to some Asian taxa

Sequence data from the nuclear ribosomal internal transcribed spacer (ITS) region for 189 accessions representing 184 species in 121 genera of Brassicaceae were used to determine monophyly of tribes and genera, tribal boundaries, and component genera. Parsimony analysis and Bayesian inference suggest that the tribes Camelineae and Arabideae are polyphyletic and should be subdivided into smaller tribes. The study also supports the recent recognition of the new tribes Aphragmeae, Biscutelleae, Buniadeae, Calepineae, Conringieae, Dontostemoneae, Erysimeae, Malcolmieae, Megacarpaeeae, and Turritideae. The data argue for the placement of Borodinia in the tribe Boechereae, Litwinowia and Pseudoclausia in the Chorisporeae, Atelanthera and Streptoloma in the Euclidieae, and Megacarpaea and Pugionium in the Megacarpaeeae, and exclusion of Asperuginoides, Didymophysa, and Ptilotrichum from the Alysseae, Macropodium, Pseudoturritis, and Stevenia from the Arabideae, and Crucihimalaya, Irenepharsus, Pachycladon, and Turritis from the Camelineae. Finally, the findings support the expansion of Stevenia to include both Berteroella and Ptilotrichum, Sterigmostemum to include Oreoloma and one species of Anchonium, Crucihimalaya to include Transberingia and several species of Arabis, and Parrya to include Pseudoclausia. The data also suggest that Calymmatium and Olimarabidopsis may be congeneric.

Molecular signals for late Tertiary/early Quaternary range splits of an Eurasian steppe plant: Clausia aprica (Brassicaceae).

Several vegetation belts stretch continuously from Europe to Asia, taiga and steppe being most prominent. Numerous plant species within these belts share a conspicuous distribution area, which is longitudinally contracted or disrupted approximately along longitude 70° E. To date no hypothesis for this intriguing distribution pattern has been put forward. We detected molecular footprints in the contemporary genetic composition in nuclear DNA (ITS1, ITS2) and chloroplast DNA (trnL–trnF spacer region) of the steppe element Clausia aprica (Brassicaceae) providing evidence for a severe longitudinal range split and genetic differentiation east of the Ural Mountains about 1 million years ago caused by Quaternary climatic oscillations. Clausia aprica provides the first phylogeographical analysis on the intraspecific evolution of an Eurasian steppe plant.

RAPDs and noncoding chloroplast DNA reveal a single origin of the cultivated Allium fistulosum from A. altaicum (Alliaceae)

The origin of the crop species Allium fistulosum (bunching onion) and its relation to its wild relative A. altaicum were surveyed with a restriction fragment length polymorphism (RFLP) analysis of five noncoding cpDNA regions and with a random amplified polymorhic DNA (RAPD) analysis of nuclear DNA. Sixteen accessions of A. altaicum, 14 accessions of A. fistulosum, representing the morphological variability of the species, and five additional outgroup species from Allium section Cepa were included in this study. The RFLP analysis detected 14 phylogenetically informative character transformations, whereas RAPD revealed 126 polymorphic fragments. Generalized parsimony, neighbor-joining analysis of genetic distances, and a principal co-ordinate analysis were able to distinguish the two species, but only RAPD data allowed clarification of the interrelationship of the two taxa. The main results of this investigation were: (1) A. fistulosum is of monophyletic origin, and (2) A. fistulosum originated from an A. altaicum progenitor, making A. altaicum a paraphyletic species. Compared with A. altaicum the cultivated accessions of the bunching onion show less genetic variability, a phenomenon that often occurs in crop species due to the severe genetic bottleneck of domestication. Allium altaicum and A. fistulosum easily hybridize when grown together, and most garden-grown material is of recent hybrid origin.

Origin of some minor vegetatively propagated Allium crops studied with RAPD and GISH

Three vegetative crops of sect. Cepa in genus Allium (Top onion, French grey shallot and viviparous triploid onion) of suspected hybridogenic origin were studied with genomic in situ hybridization (GISH) and random amplified polymorphic DNA (RAPD) markers. Related wild and cultivated species were included in the analysis in order to assess their contributions to the genomes of the investigated species. In A. × proliferum, the parental chromosomes derived from A. fistulosum and A. cepa were unequivocally identified by GISH, proving the hybrid status of this crop. The French grey shallot proved to belong to A. oschaninii according to the RAPD analysis and the GISH results, it is clearly separate from the normal shallots of A. cepa var. aggregatum. Thus the grey shallots are a new crop species and can be considered as an aggregatum form of the wild progenitor species A. oschaninii. The triploid viviparous onion comprises mostly A. cepa derived DNA in its genome. A non-cepa component could not be attributed to any of the sect. Cepa species included in this study and is most likely derived from one of the species of this section not yet identified by molecular means.

‘Missing link’ species Capsella orientalis and Capsella thracica elucidate evolution of model plant genus Capsella (Brassicaceae)

To elucidate the evolutionary history of the genus Capsella, we included the hitherto poorly known species C. orientalis and C. thracica into our studies together with C. grandiflora, C. rubella and C. bursa‐pastoris. We sequenced the ITS and four loci of noncoding cpDNA regions (trnL – F, rps16, trnH –psbA and trnQ –rps16). Sequence data were evaluated with parsimony and Bayesian analyses. Divergence time estimates were carried out with the software package BEAST. We also performed isozyme, cytological, morphological and biogeographic studies. Capsella orientalis (self‐compatible, SC; 2n = 16) forms a clade (eastern lineage) with C. bursa‐pastoris (SC; 2n = 32), which is a sister clade (western lineage) to C. grandiflora (self‐incompatible, SI; 2n = 16) and C. rubella (SC; 2n = 16). Capsella bursa‐pastoris is an autopolyploid species of multiple origin, whereas the Bulgarian endemic C. thracica (SC; 2n = 32) is allopolyploid and emerged from interspecific hybridization between C. bursa‐pastoris and C. grandiflora. The common ancestor of the two lineages was diploid and SI, and its distribution ranged from eastern Europe to central Asia, predominantly confined to steppe‐like habitats. Biogeographic dynamics during the Pleistocene caused geographic and genetic subdivisions within the common ancestor giving rise to the two extant lineages.

Phylogenetic relationships of wild and cultivated species of Allium section Cepa inferred by nuclear rDNA ITS sequence analysis

Allium section Cepa consists of 12 species most of which are used by humans as condiment, vegetable or medicinal plants. Common onion (Allium cepa) and bunching onion (A. fistulosum) are cultivated species while all others are locally collected from the wild. Although common onion is the most important crop within Allium, its wild progenitor and origin are still not clear. We analyzed the phylogeny of Allium section Cepa using sequences of the nuclear ribosomal DNA internal transcribed spacer (ITS) region of 36 accessions representing eleven species of section Cepa, together with eight outgroup species with phenetic, cladistic, and model-based algorithms. These analyses confirmed section Cepa to be monophyletic and revealed three species groups within the section. These are (i) A. altaicum/A. fistulosum, (ii) A. farctum/A. roylei/A. asarense/A. cepa/A. vavilovii, and (iii) A. galanthum/A. oschaninii/A. praemixtum/A. pskemense. While the first two groups were statistically well supported for the last group support was low, although it resulted in all phylogenetic analyses conducted. Tree and network-based analyses grouped A. cepa within A. vavilovii, indicating the latter to be progenitor of the common onion. However, also an origin of A. cepa through hybridization of A. vavilovii with A. galanthum or A. fistulosum seems possible. We argue that a subdivision of section Cepa in subsections Cepa and Phyllodolon, although possible from our data, as well as the formal description of alliances do not seem reasonable in a small group of species.

Assessment, validation and deployment strategy of a two- barcode protocol for facile genotyping of duckweed species ‡

Lemnaceae, commonly called duckweeds, comprise a diverse group of floating aquatic plants that have previously been classified into 37 species based on morphological and physiological criteria. In addition to their unique evolutionary position among angiosperms and their applications in biomonitoring, the potential of duckweeds as a novel sustainable crop for fuel and feed has recently increased interest in the study of their biodiversity and systematics. However, due to their small size and abbreviated structure, accurate typing of duckweeds based on morphology can be challenging. In the past decade, attempts to employ molecular barcoding techniques for species assignment have produced promising results; however, they have yet to be codified into a simple and quantitative protocol. A study that compiles and compares the barcode sequences within all known species of this family would help to establish the fidelity and limits of this DNA-based approach. In this work, we compared the level of conservation between over 100 strains of duckweed for two intergenic barcode sequences derived from the plastid genome. By using over 300 sequences publicly available in the NCBI database, we determined the utility of each of these two barcodes for duckweed species identification. Through sequencing of these barcodes from additional accessions, 30 of the 37 known species of duckweed could be identified with varying levels of confidence using this approach. From our analyses using this reference dataset, we also confirmed two instances where mis-assignment of species has likely occurred. Potential strategies for further improving the scope of this technology are discussed.

Allotetraploid origin ofAllium altyncolicum (Alliaceae, Allium sect.Schoenoprasum) as investigated by karyological and molecular markers

The tetraploidAllium altyncolicum (2n = 4x = 32) is considered to be of hybrid origin, because most of its morphological characters are intermediate between those of its putative parents,A. schoenoprasum andA. ledebourianum. In the present work an attempt has been made to ascertain its parentage by several methods: Giemsa C-banding, genomic in situ hybridization (GISH), PCR-RFLP of cpDNA, restriction enzyme mapping of the rDNA, and RAPDs. C-banding and GISH indicates clearly thatA. altyncolicum is a segmental allopolyploid.Allium schoenoprasum andA. ledebourianum are the most likely the parental species and the larger part of the genome ofA. altyncolicum (26 chromosomes) is derived fromA. schoenoprasum. The low genetic divergence between these three species was confirmed by the lack of sequence variation in the ITS sequences of nuclear rRNA genes and of the plastid rbcL-atpB intergenic spacer. Both parental species andA. altyncolicum could be distinguished by RFLP of the rDNA repeats. The geographic origin of the putative parental species was investigated using RAPDs.

Phylogeny of Allium L. subgenus Anguinum (G. Don. ex W.D.J. Koch) N. Friesen (Amaryllidaceae).

The disjunct distribution of the subgenus Anguinum of the genus Allium makes it a good candidate to test models of Northern hemisphere biogeography. Here we conduct phylogenetic analysis with the nuclear marker ITS and three different chloroplast markers (rps16 intron, rbcL-atpB spacer, rpl32-trnL spacer). Divergence time estimations (Beast) relying on published ITS substitution rates and ancestral range reconstructions were calculated to elucidate the biogeographical history of the subgenus. Additionally we compiled distribution maps for all species with data taken from the literature, herbariums and data from field observations. The main radiation of the subgenus took place in the last one million years and is still going on. They have their origins in the mid Miocene in East Asia and were highly influenced by the climate fluctuations in the Pliocene/Pleistocene period. Conflicting tree topologies between nuclear and cpDNA markers of Allium tricoccum Solander indicate that the species is of hybridogenous origin. Cloning the ITS sequence revealed the parental copies and confirmed our conclusion. One originated from the Eurasian and the other from the East Asian clade. We were able to show that it reached North America most likely via the Beringia around 2.5mya (95% HPD of 0.35-5.26mya). Our data suggest that Allium victorialis L. is only distributed in mountain pastures in Europe as it forms a well-supported clade in the ITS tree. In the analysis of the molecular markers we found two distinct types of Allium ochotense Prokh. and we suggest splitting the species based on Prokhanovs (1930) proposal. Taxonomical remarks and an identification key to all species of the subgenus Anguinum is provided.