Jan Kirschner

Comparison of seven DNA extraction and amplification protocols in historical herbarium specimens of juncaceae

Seven DNA extraction protocols were used to obtain DNA from herbarium specimens ofJuncus andLuzula (Juncaceae) of various ages. DNA of historical samples is difficult to extract, and the extracts are seldom of good quality. The quality of DNA obtained was estimated by using a spectrophotometer to measure the A260/280 absorbance ratio. The total DNA yield was measured by a fluorometer. The results indicate the success of using both mixer mill grinding and a DNeasy Plant Kit. Another extraction protocol (grinding with mortar and pestle, using liquid nitrogen) yielded DNA from many samples. Modified CTAB extraction, with a lengthy precipitation, usually provided good amounts of DNA. Other protocols did not give satisfactory results.

Modes of speciation and evolution of the sections inTaraxacum

Modes of evolution of species classified within different sections inTaraxacum involve diverse processes, viz. primary divergence of an ancestral sexual diploid, hybridization between a tetraploid apomict and a diploid sexual hybrid, differentiation of an advanced apomictic taxon at one ploidy level, hybridization between a sexual tetraploid and a sexual diploid, formation of a polyploid series from an apomictic ancestor of a lower polyploidy level, and remote hybridization between an autumn-flowering ancestral diploid and a spring-flowering derivative diploid or apomict. Various reproduction systems of the plants involved, different environments and different timing of the processes contribute to a very varied nature of the species groups.

A nomenclatural checklist of supraspecific names in Taraxacum

Summary legitimacy are determined, all valid names are given types, and type specimens are selected and quoted where available. Comments are confined to the nomenclatural status of the names; for important taxonomic synonyms the accepted names are given, and names considered correct are highlighted.

Introgressive hybridization between a native and an introduced species:Viola lutea subsp.Sudetica versusV. Tricolor

A hybrid swarm betweenViola lutea subsp.sudetica (2n=50, native species) andViola tricolor (2n=26, introduced species) originated in the 1970’s in the Krkonoše Mts. Analyses of chromosome numbers, reproductive biology, morphology, and ecology gave the following results: (1) Compared to the plants found in the 1970’s, the number of colour morphs have decreased and the types now prevailing in the field are morphologically closer toV. lutea subsp.sudetica, forming a continuum. (2) In the field, hybrids having approximately the same chromosome number as the primary hybrids are most common. Some plants of the hybrid swarm have certain characters unknown to their parents. (3) In the field, autogamous types similar toV. tricolor were found. (4) A limited number of plants from the field and culture have higher somatic chromosome numbers thanV. lutea subsp.sudetica; there was a tendency towards increasing chromosome numbers in their progeny (up to 2n=62). These plants have some new morphological characters (a small proportion of hexacolpate pollen) not found in the parents (nor in the other hybrids with prevailing tetracolpate and pentacolpate pollen grains) and higher pollen fertility in comparison to the other hybrids. These plants also have the highest germination rate. (5) There is a tendency for chromosome numbers to decline in the progeny of open pollinated hybrid plants in the lowland experimental graden. (6) The phenology of the plants in the mountain grasslands and the lowland garden is different; the parents behave in a totally contrasting manner. (7) In the field at least some hybrids extend outside the geographical and ecological ranges of the parental species, invading new communities. (8) Seeds ofV. tricolor do not show any dormancy and germinate in the year of production; most of theV. lutea subsp.sudetica seeds germinate during the spring of the following year. Hybrids with intermediate chromosome numbers had both types of germination strategy. The germination rate of intermediates with high chromosome numbers was even higher than that ofV. tricolor.

Phylogeny of the Juncaceae based on rbcL sequences, with special emphasis on Luzula DC. and Juncus L.

Abstract.Cladistic analysis of rbcL nucleotide sequences was applied to 58 taxa representing most subgenera and sections of Luzula and Juncus, chosen to reflect morphological and geographical diversity of both genera. Additionally, representatives of all other genera of the Juncaceae and two taxa from the Cyperaceae were included. Phylogenetic trees were constructed using parsimony with Prioniumserratum as outgroup. The dataset has 190 parsimony informative sites. The analysis yielded more than 332,400 equally parsimonious trees (length 620, CI=0.47, RI=0.82). A jackknife analysis revealed several well-supported clades. Luzula is monophyletic and Juncus is non-monophyletic. Each of the generally accepted subgenera of Juncus, subg. Juncus and subg. Agathryon, form a clade, but their circumscription differs from the traditional views. The subgenera recognized in Luzula remain mainly unresolved. A well-supported clade is represented by an assemblage of representatives of five genera and species distributed in the Southern Hemisphere: Juncuscapensis and J. lomatophyllus (both from section Graminifolii), Rostkovia, Distichia, Marsippospermum, and Patosia.

Genetic variation in agamospermous taxa of Hieracium sect. Alpina (Compositae) in the Tatry Mts. (Slovakia)

Abstract. The mode of reproduction, pollen production, chromosome numbers, genetic variation (RAPD, allozymes) and overall similarity were studied in 6 species of Hieracium sect. Alpina in the Tatry Mts. (the Western Carpathians, Slovakia). All species were confirmed to be agamospermous and, except of H. krivanense and H. slovacum, lacking pollen grains. For the first time, a chromosome number is reported for H. krivanense (2n=4x=36). Considerable genetic variation was revealed in H. alpinum and a correlation between geographic and genetic distances was found in this species. Between-population variation in RAPD and allozyme phenotypes was found in H. pinetophilum and H. crassipedipilum. In all other species, allozyme and RAPD variation was low or absent. With few exceptions, the species differ in their allozyme as well as RAPD patterns. The relatedness of one population of endemic H. slovacum and H. halleri was confirmed. It is shown, that Carpathian species of the H. fritzei group are derived from at least two ancestors.

Analysis of nrDNA polymorphism in closely related diploid sexual, tetraploid sexual and polyploid agamospermous species

Nuclear sequences of ITS1-5.8S-ITS2 region of rDNA may be an important source of phylogenetically informative data provided that nrDNA is cloned and the character of sequence variation of clones is properly analyzed. nrDNA of selected Taraxacum sections was studied to show sequence variation differences among diploid sexual, tetraploid sexual and polyploid agamospermous species. We examined nucleotide characteristics, substitution pattern, secondary structure, and the phylogenetic utility of ITS1-5.8S-ITS2 from 301 clones of 32 species representing 11 sections. The most divergent sequences of ITS1&2 differed by 17.1% and in 5.8S only by 3.7%. The ITS1-5.8S-ITS2 characteristics, integrity and also stability of secondary structures confirmed that pseudogenes are not responsible for the above variation. The within-individual polymorphism of clones implies that the concerted evolution of ITS cistron of agamospermous polyploid Taraxacum is remarkably suppressed. Sequences of ITS clones proved to be a useful tool for mapping pathways of complex reticulation (polyploid hybridity) in agamospermous Taraxacum.

Variation inTaraxacum bessarabicum and allied taxa of the sectionPiesis (Compositae): allozyme diversity, karyotypes and breeding behaviour

Allozyme techniques, karyotype analyses and cultivation experiments were carried out on 20 population samples ofTaraxacum sect.Piesis (Compositae), viz.T. bessarabicum (17 samples from W. and C. Europe, Ukraine and Crimea, Central Asia and the Altai),T. salsum from Crimea,T. x mesohalobium from Crimea, andT. stenolepium from the Causasus. The taxa studied share a primitive, symmetrical karyotype. All taxa studied are sexual,T. bessarabicum mostly autogamous. Sexuality at the tetraploid level (T. stenolepium) and occasional male sterility in a sexual (T. bessarabicum in Moravia) were recorded for the first time in the genus. Selfing, diploidy and the highly predictable habitat may account for the mostly low level within population allozyme variation inT. bessarabicum. W. and C. European samples ofT. bessarabicum are almost invariable allozymically and, as a group, have no unique alleles. The Crimean and Asian group of populations shows higher levels of allozyme variation and has 15 alleles not found in the former group at 13 loci studied. Recent migration from one source region is suggested to account for the homogeneity of the western group, refugial persistence of alleles and possible introgression from sympatric species may have resulted in allele richness in the eastern group. Hybridization betweenT. bessarabicum andT. salsum was documented by allozyme patterns in a few plants in Crimea. Some aberrant allozyme or karyotypic features of two populations are discussed as well.

A revision ofTaraxacum sect.Piesis (Compositae)

On the basis of allozyme and cultivation data, and of additional herbarium material, a taxonomic and nomenclatural revision ofTaraxacum sect.PiesisA.J. Richards exKirschner etŠtěpánek is provided. The section is made up of halophilous, sexually reproducing taxa. InT. stenocephalumBoiss. etKotschy,T. pindicumKirschner etŠtěpánek, sp. nov., andT. perenneKirschner etŠtěpánek, sp. nov., a tetraploid chromosome number has been recorded, representing the only known case of sexuality at the tetraploid level in the genus. The complex ofT. stenocephalum, includes some geographically and morphologically extreme populations treated as subspecies: subsp.gumusanicum (Soest)Kirschner etŠtěpánek, comb. nov., subsp.magnumKirschner etŠtěpánek, subsp. nov., and subsp.daralagesicum (Schischk.)Kirschner etŠtěpánek, comb. nov. In addition toT. bessarabicum (Hornem.)Hand.-Mazz., a widely distributed Eurasian species,T. stenocephalum, a complex centred in Transcaucasia and Anatolia, andT. pachypodumH. Lindb., a North African endemic, four new species are described:T. salsumKirschner etŠtěpánek, sp. nov., a diploid endemic confined to E Crimea,T. perenneKirschner etŠtěpánek, sp. nov., a tetraploid sexual species known only from SW Crimea,T. pindicumKirschner etŠtěpánek, sp. nov., a remarkable tetraploid endemic to the Pindos Mts., Greece, andT. salsitatisKirschner, Štěpänek etYirdirimli, sp. nov., an Anatolian diploid species. Furthermore, a hybrid betweenT. salsum andT. bessarabicum from Crimea (documented on the basis of allozyme data elsewhere) is given a binomial,T. xmesohalobiumKirschner etŠtěpánek, nothosp. nov.

New sections inTaraxacum

Sectional taxonomy ofTaraxacum in steppe or subsaline habitats in Central Asia is revised based on material collected during expeditions, cultivated or studied in herbarium. Two new sections are described from that area:T. sect.Stenoloba similar toT. sect.Leucantha (syn.:T. sect.Sinensia), andT. sect.Suavia allied toT. sect.Dissecta. The type species of the sectionSuavia is described asTaraxacum formosissimumKirschner etŠtěpánek. Widespread mountain dandelions of the Caucasus, intermediate between the sect.Piesis andT. stevenii, are described asT. sect.Confusa. Taraxacum species dominating dry habitats in S Ukraine and Crimea are described asT. sect.Borysthenica. Species belonging to the new sections were found to be polyploid and agamospermous.