Biometry and DNA fingerprinting detect limited genetic differentiation among populations of the apomictic blackberry Rubus nessensis (Rosaceae)

Clonal plant species have in many recent studies exhibited unexpectedly high levels of genetic differentiation. This seems to hold true also for several apomictic (agamospermous) species in spite of their potential ability to spread single genotypes over large areas through seed dispersal. By contrast, previous DNA fingerprint studies have demonstrated apomictic Rubus species to be extremely homogeneous. The present study was undertaken to estimate the extent and distribution of intraspecific genetic variation in R. nessensis . Populations differed significantly for many biometrical characters but this variation was evenly distributed, with few significant differences between single pairs of populations. Patterns indicating ecoclinal variation, which could be adaptive in origin, were not found. Populations also clustered close together in a two-dimensional CVA plot, with only one population being somewhat further removed. DNA analysis of later collected material from this population as well as from three others yielded identical DNA fingerprints with each of two different multi-locus DNA probes. Taken together with previous DNA fingerprint studies, these results indicate that a major part of the R. nessensis plants in Northern Europe have been derived from a single recombinational event, possibly making this genet the most widespread land-living organism in the world. The encountered variation in biometrical characters is probably instead caused by e.g. minor somatic mutations. Rubus species have relatively large, bird-dispersed seeds that probably travel further than the small, wind-dispersed seeds found in most previously investigated apomicts (e.g. in Asteraceae). The potential for single genotypes to disperse over large areas should therefore be greater in Rubus than in many other apomictic species groups.     

DNA fingerprint variation in some blackberry species (Rubus subg.Rubus,Rosaceae)

We have analysed samples from Sweden, Denmark, and Germany of six facultatively apomictic blackberry species to investigate the accordance between a taxonomy based on morphological characters on the one hand, and distribution of genetic variation estimated by DNA fingerprinting on the other hand. DNA fingerprint variation was found to be quite restricted in all species investigated. The first taxonomic group included three species related toRubus nessensis, two being characterized by one very widespread DNA fingerprint in all three countries and a few rare ones, whereas the third species differed between Sweden and Germany. The second taxonomic group included species related toR. gracilis. Two of these species exhibited very similar DNA fingerprints, whereas the third species deviated clearly. The utilization of DNA fingerprinting as a tool in taxonomy is discussed; most likely this method could become a useful complement to morphology, especially in plant groups with reduced levels of genetic recombination.     

Chloroplast DNA diversity in the genusRubus (Rosaceae) revealed by Southern hybridization

The variability in chloroplast DNA type of 20Rubus genotypes was examined by Southern hybridization. DNA extracted from theRubus accessions was digested with two restriction enzymes (EcoRI and EcoRV) and heterologous chloroplast DNA sequences from barley and pea were used as probes to detectRubus chloroplast DNA sequences on Southern blots ofRubus total DNA. Restriction fragment length polymorphism was detected and a total of 92 restriction fragments were generated by the probe/enzyme combinations examined. Cladistic principles based on the parsimony assumption were used to assemble a phylogenetic tree based on chloroplast restriction fragment length data. The phylogenetic tree grouped the taxonomically defined species and is in general agreement with information based on morphological criteria. However, the Japanese red raspberryR. illecebrosus was shown to have diverged considerably in terms of evolutionary time from other species in subg.Idaeobatus. Furthermore, the molecular approach provides a quantitative estimate of the relationship between species that is difficult to obtain from morphological data. In order to complement the chloroplast DNA information a ribosomal DNA probe was also included in the analysis and provided further information on the phylogenetic relationships withinRubus.     

Active self-pollination in blackberries (Rubus subgen. Rubus, Rosaceae)

Most blackberry species are apomictic but pseudogamous, i.e. embryo development is dependent on prior pollination. Two different types of stamen arrangement were noted in senescing flowers of 18 Swedish species and 2 common cultivars. The stamens of 'cross-pollinating' species seldom touch the stigmas while those of 'self-pollinating' species instead arch over the pistils and dry up in this position. All species set seed well after artificial self-pollination and are thus self-compatible. Isolation experiments were carried out during 3 years. When inflorescences were bagged, seed set was drastically lowered in 'cross-pollinators' but remained almost unaltered in 'self-pollinators'. The different responses to isolation are probably due to the inherent stamen behaviour.     

Apomixis versus sexuality in blackberries (Rubus subgen.Rubus,Rosaceae)

InRubus L. a connection seems to exist between the degree of meiotic disturbances on the one hand, and the production of unreduced embryo sacs, pollen fertility and relative seed set on the other hand. Severe meiotic disturbances commonly encountered in apomictic taxa decrease pollen fertility and thereby seed set since pollen is necessary for endosperm development. By contrast interspecific hybrids between apomictic taxa appear to be sexual and exhibit high pollen fertilities, probably due to an improved meiosis. Thus, apomixis leads to a decreased fertility inRubus, not the opposite, as often discussed.     

Chromosome numbers of European blackberries (Rubus subg.Rubus,Rosaceae)

Chromosome numbers are presented for 32 collections of 29 European blackberry species (Rubus subg.Rubus) from Germany. One species is triploid (2n = 21), 27 species are tetraploid, (2n = 28), and one species is pentaploid (2n = 35). Chromosome numbers are reported for the first time ofR. adspersus, R. amisiensis, R. calvus, R. conothyrsoides, R. contractipes, R. demissus, R. elegantispinosus, R. ferocior, R. foliosus, R. hypomalacus, R. leucandrus, R. nemorosus, R. platyacanthus, R. praecox, R. rhombifolius, andR. rhytidophyllus. Chromosome numbers forR. dasyphyllus, R. gelertii, R. glandithyrsos, R. lamprocaulos, R. lindebergii, R. macrophyllus, R. montanus, R. muenteri, R. pedemontanus, R. polyanthemus, R. senticosus, R. silvaticus, andR. vigorosus are confirmed.     

Genetic diversity inChaenomeles (Rosaceae) revealed by RAPD analysis

Genetic relatedness inChaenomeles was studied by RAPD analysis in 42 plants representing accessions of three wild species and one hybrid taxon. Amplification with 17 primers yielded a total of 156 polymorphic RAPD bands. Estimates of genetic relatedness suggest thatC. cathayensis andC. japonica are the most distantly related species, and that the former is comparatively homogeneous.Chaenomeles speciosa, which may have arisen through hybridization betweenC. cathayensis andC. japonica, takes an intermediate position between these two species. Analysis of diagnostic bands demonstrate that neitherC. speciosa norC. ×superba has any bands that do not occur in at least one ofC. cathayensis orC. japonica. Moreover,C. speciosa and the partly overlapping taxonC. ×superba are comparatively heterogeneous, which is also in accordance with a hybrid origin. Intraspecific variation was studied mainly inC. japonica; plants obtained from different sources of material formed well separated groups in the cluster analysis.     

钻地风的三萜类成分

从钻地风根的乙醇提取物中分离得到12个三萜及三萜皂苷类化合物,通过波谱分析并与有关对照品比较, 其结构鉴定为熊果酸(1),坡模酸(2),tormentic acid (3),蔷薇酸(4),sericic acid (5),23-hydroxytormentic acid (6), kaji-ichigoside F1 (7),野蔷薇苷 (8), sericoside (9), quadranoside VIII (10), crataegioside (11)和niga-ichigoside F1 (12).初步研究结果表明,这些化合物在体外5?mg/ml浓度下对耐药金黄色葡萄球菌(MRSA)无抑制作用.     

Rediscoveries of two species of Rubus (Rosaceae) in China

报道了蔷薇科两种悬钩子属植物在中国的重新发现,分别为美叶悬钩子(Rubus calophyllus C.B.Clark)和拟针刺悬钩子(R.parapungens H.Hara),并提供了简要的描述和标本照片.     

Pollination ecology of the high latitude, dioecious cloudberry (Rubus chamaemorus; Rosaceae)

In a 3-yr study, we examined the pollinator guild and intersexual floral characteristics of the dioecious, perennial cloudberry (Rubus chamaemorus), which flowers in early spring. The findings contribute to our general understanding of pollination ecology at high latitudes and provide important information for the commercialization of cloudberry. Female flowers were smaller than males but provided more nectar, although this resource was low in both sexes. Insects from 43 families visited cloudberry flowers, yet four families (Apidae, Halictidae, Muscidae, Syrphidae) represented ca. 87% of all visitors observed. Introduction experiments revealed that apids and muscids are significantly poorer pollinators (based on fruit production) than halictids and syrphids, but when fruit mass or seed set was considered, there were no significant differences between families. Pollinator importance, a product of flower visitation frequency and seed set effectiveness, revealed that the dipterans were of paramount importance to the pollination of cloudberry. Furthermore, they are limited to cloudberry because their lapping mouthparts exclude them from accessing the nutritional rewards of competing Ericaceae flowers. While the total number of pollinator families observed suggest a generalist pollination system, if one considers the dominant pollinators (flies) as a functional group, then this insect-flower relationship could be considered a specialized one.     

Species relationships in Fagopyrum revealed by PCR-based DNA fingerprinting

Random amplified polymorphic DNA (RAPD) markers were used to distinguish between 28 different accessions belonging to 14 species and two sub-species of Fagopyrum. Of the 75 random 10-mer primers tested, only 19 generated robust, easily interpretable amplification products. A total of 364 bands were observed with an average of 19.15 bands per primer, of which 99.45% were polymorphic. Primer OPN-08 produced the maximum number of fragments and UBC-183 produced the minimum number of fragments. The data were utilized to elucidate genetic relationships among 14 species and two sub-species of Fagopyrum. Cluster analysis using the unweighted paired group method of arithmetic means (UPGMA) showed four main clusters, two each of the cymosum and urophyllum groups. The results showed that Fagopyrum tataricum is closer to its wild ancestor F. tataricum ssp. potanini Batalin, closely followed by Fagopyrum giganteum. Cultivated common buckwheat ( Fagopyrum esculentum) showed affinity with its putative wild ancestor F. esculentum ssp. ancestrale and the other closely related diploid species Fagopyrum homotropicum. In the urophyllum group, Fagopyrum macrocarpum and Fagopyrum pleioramosum formed one cluster, whereas Fagopyrum capillatum, Fagopyrum gracilipes and Fagopyrum gilessii clustered separately. Except for a few cases, our results correspond with previously reported studies on Fagopyrum using the isozyme, RFLP and RAPD methods. Species-diagnostic amplification products specific to some species in the cymosum and urophyllum groups were identified. Our results show that RAPDs can be successfully used to analyze species relationships in Fagopyrum and also for constructing linkage maps.     

Photosynthetic characteristics of invasive and noninvasive species of Rubus (Rosaceae)

The prolific amount of growth and reproduction in invasive plants may be achieved by greater net photosynthesis and/or resource-use efficiency. I tested the hypotheses that leaf-level photosynthetic capacity and resource-use efficiency were greater in two invasive species of Rubus as compared with two noninvasive species that have overlapping distributions in the Pacific Northwest. The invasive species had significantly higher photosynthetic capacity and maintained net photosynthesis (A) over a longer period of the year than the noninvasive species. The construction cost (CC) of leaf tissue per unit leaf mass was comparable among the four species, but the invasive species allocated less nitrogen (N) per unit leaf mass. On a leaf area basis, both leaf CC and N were higher for the invasive species. The specific leaf area (SLA) was also lower in the invasive species, indicating less photosynthetic area per gram leaf tissue. The invasive species achieved high A at lower resource investments than the noninvasive species, including having higher maximum photosynthetic rate (A(max)) per unit dark respiration (R(d)), greater A(max) per unit leaf N (photosynthetic nitrogen-use efficiency), and greater water-use efficiency as measured by instantaneous rates of A per unit transpiration (A/E) and by integrated A/E inferred from stable carbon isotope ratios (δ(13)C). Using discriminant analysis, these photosynthetic characteristics were found to be powerful in distinguishing between the invasive and noninvasive Rubus. A(max) and A/E were identified as the most useful variables for distinguishing between the species, and therefore, may be important factors contributing to the success of these invasive species.     

The origin of the apple subfamily (Maloideae; Rosaceae) is clarified by DNA sequence data from duplicated GBSSI genes

For 70 yr the leading hypothesis for the origin of the Maloideae has involved wide hybridization between ancestors of two other subfamilies. The basis of this hypothesis is that Maloideae have a base chromosome number of 17, whereas other Rosaceae are mostly x = 7, 8, or 9. To investigate this hypothesis we cloned and sequenced approximately 1.8 kilobases from the 5' portion of granule-bound starch synthase (GBSSI, or waxy) genes for 89 clones from 32 Rosaceae genera. Previous studies demonstrate the presence of two copies in all Rosaceae (GBSSI-1 and GBSSI-2) and four in Maloideae (GBSSI-1A, GBSSI-1B, GBSSI-2A, and GBSSI-2B). Parsimony and maximum likelihood analyses nest Gillenia, a genus of the southeastern United States with a base chromosome number of 9, within either Maloideae GBSSI-1 or GBSSI-2. Monophyly of Maloideae plus Gillenia is well supported by bootstrap values, loss of the sixth intron in all GBSSI-1 sequences, intron alignability between genera, and numerous nonmolecular characters. Our results falsify the wide-hybridization hypothesis and are consistent with a polyploid origin involving only members of a lineage that contained the ancestors of Gillenia. Under this hypothesis, the subfamily originated in North America, and the high Maloideae chromosome number arose via aneuploidy from x = 18.     

Pollen stainability variation in the apomictic blackberry, Rubus nessensis (Rosaceae)

Pollen quality was assessed with cotton blue staining in an apomictic blackberry, Rubus nessensis W. Hall, on plants grown outdoors in a random arrangement under uniform conditions. Significant variation between populations may be attributed mainly to genetical factors. Most of the variation found between plants, sometimes even between ramets, within a population must, however, be environmentally caused, likewise the variation between flowers within the same plant.     

Cytotoxic,hypoglycemic activity and phytochemical analysis of Rubus imperialis (Rosaceae)

Rubus imperialis, Artemia salina, 3-O-methylellagic-4'-O-alpha-rhamnose Acid Screening of different extracts, fractions and compounds from Rubus imperialis Chum. Schl. (Rosaceae) has been conduced using the brine shrimp microwell cytotoxicity assay. Three parts of the plant (methanolic extract from leaves, roots and stems), three fractions from roots (hexane, ethyl acetate and butanol) and three isolated compounds (niga-ichigoside F1, 23-hydroxytormentic acid, ellagic acid derivative) were tested. The most promising material (LC50 <1000 microg/ml) were the methanolic extract and ethyl acetate fraction from roots. However, there was little correlation observed in the degree of toxicities observed between the isolated compounds. On the other hand, the cytotoxicity and in vivo assays confirmed the hypoglycemic activity of methanolic extract and validated the Brazilian popular use of R. imperialis as an antidiabetic agent.     

A demographic study of the apomictic blackberry, Rubus nessensis (Rosaceae)

Ten stands of Rubus nessensis , an apomictic blackberry, were studied. Perennial stools produce biennial shoots that flower in their second year. The stools tend to produce shoots of similar height for several consecutive years. Flowering, but not relative fruit set, is dependent on shoot height. Shoot height and density appear to be positively related, implying that intraspecific competition is of little importance for plant vigour. Depletion time varies greatly between stands (4–29 years), being comparatively longer in stands with short shoots. New establishment is sparse and takes place mostly by root suckering.     

Low genetic variability in a natural alpine marmot population (Marmota marmota, Sciuridae) revealed by DNA fingerprinting

Genetic heterogeneity is usually considered an important factor for the viability of a population, yet there are cases in which populations sustain themselves despite virtual homozygosity. A prior step to studying the effects of such low levels of genetic variability can be the analysis of its causes. We analysed a population of the highly social alpine marmot ( Marmota marmota , Sciuridae) by multilocus DNA fingerprinting. The fingerprint patterns revealed a very low degree of polymorphism in our main study population. We show that this lack of hypervariability is caused by a low effective population size, rather than by an unusual low mutation rate of the fingerprint loci studied. However, the current number of breeding pairs was found to be about an order of magnitude larger than the one that would be expected to lead to such a low degree of heterozygosity. We conclude that there must have been bottlenecks in the history of the Berchtesgaden marmot population that have severely affected its genetic heterozygosity.     

Low frequency of extra-pair paternity in Common Gulls (Larus canus) as revealed by DNA fingerprinting

Summary During the last decade, the mating systems of many bird species have been analysed using molecular genetic methods. Most of these studies were performed on Passeriformes while Non-Passeriformes were rarely investigated. To fill this gap, we analysed blood samples of 24 Common Gull (Larus canus) families from two Polish colonies using DNA fingerprinting. A total of 55 of 56 chicks was unequivocally assigned to their genetic parents. Only two of these chicks (3.6%) from two different broods resulted from extra-pair copulations, the other 53 could be attributed to the putative parents; i.e., intraspecific brood parasitism was not observed. With respect to the number of breeding pairs investigated, the frequency of extra-pair paternity amounted to 8.3%. Thus, extra-pair paternity is relatively infrequent compared to other bird species. This result is surprising because the percentage of observed extra-pair copulations (i.e., mainly forced copulations in Common Gulls) in relation to the absolute number of copulations was 15.7% in the colonies studied. A possible reason for the disproportionate number of forced copulations and the lower percentage of resulting extra-pair young may include different mechanisms for paternity control by the pair males and/or by females.

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Niedriger Anteil von Vaterschaften außerhalb des Paarbundes bei Sturmmöwen (Larus canus)

Zusammenfassung In den letzten 10 Jahren wurde das Paarungssystem vieler Vogelarten auch mit Hilfe molekulargenetischer Methoden näher untersucht. Dabei standen vor allem verschiedene Singvogelarten im Vordergrund, während Nicht-Singvogelarten kaum berücksichtigt wurden. Die vorliegende Untersuchung an Sturmmöwen (Larus canus) soll helfen diese Lücke zu schließen. Hierzu wurden die Blutproben von 24 Sturmmöwenfamilien zweier polnischer Kolonien mit Hilfe des genetischen Fingerabdruckverfahrens untersucht. Bei 55 der 56 untersuchten Küken konnten eindeutige Aussagen zur Elternschaft gemacht werden. Nur zwei Küken (3,6%) aus zwei unterschiedlichen Bruten gingen auf Kopulationen außerhalb des Paarbundes zurück, die restlichen 53 stammten von den sozialen Eltern ab, d. h. Fälle von innerartlichem Brutparasitismus konnten nicht nachgewiesen werden. In bezug auf den prozentualen Anteil an Brutpaaren, deren Bruten Jungtiere enthielten, die das Resultat von Kopulationen außerhalb des Paarbundes waren, ergab sich ein Wert von 8,3%. Dieser Wert ist im Vergleich mit anderen Vogelarten sehr niedrig. Der geringe Anteil an Vaterschaften außerhalb des Paarbundes verwundert, da der Anteil von Kopulationen außerhalb des Paarbundes, die von den Männchen bei Sturmmöwen meist gewaltsam erzwungen werden, im Vergleich zur Gesamtzahl beobachteter Kopulationen in den untersuchten Kolonien 15,7% betrug und damit deutlich höher lag, als dies auf der Basis der Ergebnisse der DNA-Fingerprinting Untersuchungen zu erwarten gewesen wäre. Mögliche Gründe für die Diskrepanz zwischen dem Anteil der erzwungenen Kopulationen außerhalb des Paarbundes und dem Anteil der resultierenden Küken sind verschiedene Mechanismen der Vaterschaftskontrolle durch die Paarmännchen und/oder durch die Weibchen.