Duckweeds are a group of monocotyledonous aquatic plants in the Araceae superfamily, represented by 37 species divided into five genera. Duckweeds are the fastest growing flowering plants and are distributed around the globe; moreover, these plants have multiple applications, including biomass production, wastewater remediation, and making pharmaceutical proteins. Dotted duckweed (Landoltia punctata), the sole species in genus Landoltia, is one of the most resilient duckweed species. The ribosomal DNA (rDNA) encodes the RNA components of ribosomes and represents a significant part of plant genomes but has not been comprehensively studied in duckweeds. Here, we characterized the 5S rDNA genes in L. punctata by cloning and sequencing 25 PCR fragments containing the 5S rDNA repeats. No length variation was detected in the 5S rDNA gene sequence, whereas the nontranscribed spacer (NTS) varied from 151 to 524 bp. The NTS variants were grouped into two major classes, which differed both in nucleotide sequence and the type and arrangement of the spacer subrepeats. The dominant class I NTS, with a characteristic 12-bp TC-rich sequence present in 3–18 copies, was classified into four subclasses, whereas the minor class II NTS, with shorter, 9-bp nucleotide repeats, was represented by two identical sequences. In addition to these diverse subrepeats, class I and class II NTSs differed in their representation of cis-elements and the patterns of predicted G-quadruplex structures, which may influence the transcription of the 5S rDNA. Similar to related duckweed species in the genus Spirodela, L. punctata has a relatively low rDNA copy number, but in contrast to Spirodela and the majority of other plants, the arrangement of the 5S rDNA units demonstrated an unusual, heterogeneous pattern in L. punctata, as revealed by analyzing clones containing double 5S rDNA neighboring units. Our findings may further stimulate the research on the evolution of the plant rDNA and discussion of the molecular forces driving homogenization of rDNA repeats in concerted evolution.

Duckweeds represent a small, free-floating aquatic family (Lemnaceae) of the monocot order Alismatales with the fastest growth rate among flowering plants. They comprise five genera (Spirodela, Landoltia, Lemna, Wolffiella, and Wolffia) varying in genome size and chromosome number. Spirodela polyrhiza had the first sequenced duckweed genome. Cytogenetic maps are available for both species of the genus Spirodela (S. polyrhiza and S. intermedia). However, elucidation of chromosome homeology and evolutionary chromosome rearrangements by cross-FISH using Spirodela BAC probes to species of other duckweed genera has not been successful so far. We investigated the potential of chromosome-specific oligo-FISH probes to address these topics. We designed oligo-FISH probes specific for one S. intermedia and one S. polyrhiza chromosome (Fig. 1a). Our results show that these oligo-probes cross-hybridize with the homeologous regions of the other congeneric species, but are not suitable to uncover chromosomal homeology across duckweeds genera. This is most likely due to too low sequence similarity between the investigated genera and/or too low probe density on the target genomes. Finally, we suggest genus-specific design of oligo-probes to elucidate chromosome evolution across duckweed genera.

The monophyletic duckweeds comprising five genera within the monocot order Alismatales are neotenic, free-floating, aquatic organisms with fast vegetative propagation. Some species are considered for efficient biomass production, for life stock feeding, and for (simultaneous) wastewater phytoremediation. The ancestral genus Spirodela consists of only two species, Spirodela polyrhiza and Spirodela intermedia, both with a similar small genome (~160Mbp/1C). Reference genome drafts and a physical map of 96 BACs on the 20 chromosome pairs of S. polyrhiza strain 7498 are available and provide useful tools for further evolutionary studies within and between duckweed genera. Here we applied sequential comparative multicolor fluorescence in situ hybridization (mcFISH) to address homeologous chromosomes in S. intermedia (2n=36), to detect chromosome rearrangements between both species and to elucidate the mechanisms which may have led to the chromosome number alteration after their evolutionary separation. Ten chromosome pairs proved to be conserved between S. polyrhiza and S. intermedia, the remaining ones experienced, depending on the assumed direction of evolution, translocations, inversion, and fissions, respectively. These results represent a first step to unravel karyotype evolution among duckweeds and are anchor points for future genome assembly of S. intermedia.

Duckweeds, the fastest growing angiosperms, are gaining increasing attention with respect to their practical applications. Different clones of the same duckweed species vary in their physiological properties. Hence, screening of suitable clones of a species is very important. To enable the identification of clones, a clear taxonomic classification and barcoding at different taxonomic levels, i.e. genera, species, and clones is a pre-requisite. In the present project, we have focused on the genera Spirodela and Landoltia. Spirodela polyrhiza (L.) Schleid. (42 clones), Spirodela intermedia W. Koch (14 clones), and Landoltia punctata (G. Meyer) Les & Crawford (15 clones) were characterized using three plastidic sequences (rpl16, rps16, atpF-atpH) and AFLP fingerprinting. Genome size determination showed significant differences between the two genera. The genetic variability is lowest in S.polyrhiza and highest in S.intermedia. Although the resolution of phenetic variability by AFLP fingerprinting is much higher than the sequence variation of the selected plastidic regions, not all clones could be identified unequivocally. However, without any exception, all clones were strictly categorized into the three species as defined by the morphological markers. The results do not justify the separation of some clones as Spirodela biperforata from S. intermedia.

A recent (2011) attempt to change the previously designated type of the name of a duckweed species is discussed. Lemna punctata was first applied by Meyer in 1818 to a plant collected from South America, but original specimens have not been located. A prior neotype designation associated this name with a species native to parts of Asia, Australia and the Pacific, and widely introduced elsewhere, including South America. The species is generally treated by taxonomists in the genus Spirodela (either as S. punctata or the synonym S. oligorrhiza) or, more recently, as the sole member of the new (1999) genus Landoltia (as L. punctata). If accepted, this 2011 attempt to re-neotypify L. punctata would disrupt the names of two duckweed species as well as that of Landoltia. Nomenclatural arguments against accepting this new typification are provided, thereby supporting the continued usage of Landoltia in the sense intended by its original authors.

Duckweed is widely used in environmental biotechnology and has recently emerged as a potential feedstock for biofuels due to its high growth rate and starch content. The genetic diversity and composition of a natural duckweed population in genera Spirodela, Landoltia and Lemna from Lake Tai, China, were investigated using probabilistic analysis of multilocus sequence typing (MLST). The 78 strains were categorized into five lineages, among which strains representing L. aequinoctialis and S. polyrhiza were predominant. Among the five lineages, interlineage transfers of markers were infrequent and no recombination was statistically detected. Tajima's D tests determined that all loci are subject to population bottlenecks, which is likely one of the main reasons for the low genetic diversity observed within the lineages. Interestingly, strains of L. turionifera are found to contain small admixture from L. minor, providing rare evidence of transfer of genetic materials in duckweed. This was discussed with respect to the hypothesis that a cross of these two gave rise to L. japonica. Moreover, the conventional maximum-likelihood phylogenetic analysis clearly recognized all the species in the three genera with high bootstrap supports. In conclusion, this work offers a basic framework for using MLST to characterize Spirodela, Landoltia and in particular Lemna strains at the species level, and to study population genetics and evolution history of natural duckweed populations.

In this study two genera Spirodela Schleiden, and Landoltia Les & Crawford were recorded for the first time for flora of Iraq. Each genus represented by one species, these species is Spirodela polyrhiza (L.) Schleiden and Landoltia punctata (G. Meyer) Les & Crawford. In addition one more species of the genus Lemna (L. minuta) was added to the flora of Iraq for the first time too. Brief descriptions with main distinguishing characters for the three new recorded species were provided. Habitat and geographical distribution were mentioned .Floral and fruit characters for Lemnaceae species from Iraqi material were tabulated and illustrated for the first time. A key to all species of Lemnaceae in Iraq was provided.

The trnL-trnF spacer region of the chloroplast tRNA genes was sequenced and characterized in 14 accessions of the genus Spirodela (Lemnaceae). Only a low intraspecific variation of the spacer was observed in geographically isolated and morphologically different accessions of S. polyrrhiza, the most widespread Spirodela species. Five haplotypes of the spacer were identified, differing in mono-and oligonucleotide repeats and extended indels, specific to S. polyrrhiza, Landoltia punctata, and Lemna sp. The result supported the isolation of Landoltia from Spirodela.

Several photosystem II (PSII) proteins - LHCII, D1, D2, CP43 - are known to undergo light dependent reversible phosphorylation. However, little is known about the kinases responsible for their phosphorylation. Using as substrate a synthetic peptide mimicking the N-terminus of D1, we have isolated, identified and characterized a thylakoidal protein kinase. The putative D1 kinase is an extrinsic membrane protein of 50 kD with an optimum pH of 7.4, isoelectric point of 5.5, and preference for manganese cations. The N-terminus of the purified protein was blocked. Therefore, several tryptic fragments of the protein were sequenced, based on which we synthesized degenerate oligonucleotide primers to clone the gene from rice, Arabidopsis and Spirodela. The cDNA sequence predicts a protein kinase with a mass of 59 kD based on the entire open reading frame. This size is compatible with processing of a primary translation product to produce a mature protein like that recovered from purification of the rice kinase. The protein kinase gene identified shows sequence similarities to members of the family of calcium dependent protein kinases (CDPK?s; Breviario et al. Plant Mol. Biol. 27: 953-967, 1995). One of the CDPK isoforms from Spirodela but not the rice or Arabidopsis CDPK is imported into intact pea chloroplasts, although at a significantly slower rate than pLHCII. Spirodela kinase expressed in E. coli as an N-terminal 6-His fusion protein was purified. The purified kinase phosphorylated the N-terminal synthetic D1 peptide, but not LHCII or Rubisco peptides. Work is in progress on the role of CDPK in chloroplast function.

Flowering in the genus Spirodela, whether in the laboratory or in nature, has been observed only rarely. In this communication, the growth and flowering behaviour of a local isolate of S. polyrrhiza, strain SP 20 , is being reported. The presence of a chelate, such as EDTA, is obligatory for satisfactory vegetative growth of S. polyrrhiza SP 20 - An optimal flowering response is obtained, however, only when compounds such as EDDHA, a phenolic analog of EDTA, or benzoic acid are supplied. Flowering, so induced, is not influenced by the length of the photoperiod. Flowering fronds become gibbous and both EDDHA and benzoic acid also enhanced anthocyanin content. This investigation has also revealed that salicylic acid, which is known to have chelating properties itself, induces flowering in this duckweed only in the simultaneous presence of EDTA, in the nutrient solution.

Urease in Spirodela.

This is a study of the small animals in an association of one of the smallest flowering plants, Lemna minor, in the vicinity of Ithaca, N. Y. The dominant animal forms are naturally very small, ranging in size from a fraction of a millimeter to scarcely six millimeters in length. Yet, just as with large organisms, the destruction caused by them is compensated by the prolific growth of the plant. The individual plants of Lemna mttinor are, as is well known, very small, flat, leaf-like discs, convex on the upper side and slightly concave underneath. A single water root dangles from the center of each green thallus and helps to maintain the equilibrium of the plant. Being a free-swimming hydrophyte, Lemn1a minor has many air cavities which make it buoyant. Stomata are scattered over the upper surface. From April to October may be considered the active growing period of this species in the latitude of Ithaca, New York, with the peak of rapidity in growth occurring in July and August, when an entire pond may be blanketed with these small plants. Proliferation takes place in one plane, the lateral branches emerging from two clefts or pouches on the margin of the thallus. These new thalli are connected with the parent thallus by short slender stalks and are easily detached by a slight disturbance of the water. New centers of growth are thus established in the pond by the drifting apart of the disconnected offspring. In the fall, when conditions become less favorable for growth, lateral branches develop which lack air spaces and have plenty of reserve food. These winter buds sink to the bottom of the pond where they remain until the water becomes warm again the next spring. Then with the formation of air spaces, they begin their vegetative activities for the summer and rise to the surface. Propagation takes place also by the production of seeds from diminutive flowers (fig. 1). Inasmuch as Leminia mi-nor grows in quiet waters, it is not surprising that many filamentous forms of algae, as well as colonial types, are found among its roots and attached to the under surface of the thalli. Desmids and diatoms likewise abound. The liverwort, Riccia natans, and the spermatophytes, Spir odela and Wolfia, sometimes float about with the Lem-na. Near the edges of ponds grow cat-tails (Typhaceae), bur-reeds (Sparganiaceae), and arrowheads (Alismaceae). Occasionally water-lilies (Nymphaeaceae), water cress (Cruciferae), Elodea, and CeratophyllvUn associate themselves with the duckweeds (Lemlnaceae).