HAPLOID PARTHENOGENETIC SPOROPHYTES OF LAMINARIA JAPONICA (PHAEOPHYCEAE)1

Parthenogenetic sporophytes were obtained from three strains of Laminaria japonica Areschoug. These sporophytes grew to maturity in the sea, producine spores that all grew into female gametophytes. These female gametophytes gave rise to another generation of parthenogenetic sporophytes during the next year, so that by the year 1990 parthenogenetic sporophytes had been cultivated for 12, 9, and 7 generations, respectively, for the three strains. When female gametophytes from parthenogenetic sporophytes were combined with normal male gametophytes, normal sporophytes that reproduced and gave rise to both female and male gametophytes were obtained. The parthenogenetic sporophytes were shorter and narrower than the normal sporophytes of the same strain. Chromosome counts on mature sporophytes showed that normal sporophytes (from fertilized eggs) were diploid (2n = approximately 40) and that the spores they produced were haploid (n = approximately 20), while nuclei from both somatic and sporangial cells in parthenogenetic sporophytes were haploid. All gametophytes were haploid. Young sporophytes derived from cultures with both female and male gametophytes were diploid, while young, sporophytes obtained from female gametophytes from parthenogenetic sporophytes had haploid, diploid, or polyploidy chromosome numbers. Polyploidy was associated with abnormal cell shapes. The presence of haploid parthenogenetic sporophytes should be use in breeding kelp strains with useful characteristics, since the sporophyte phenotype is expressed from a haploid genotype which can be more readily selected.     

Mechanisms governing sex-ratio variation in dioecious Rumex nivalis

Sex ratios of flowering individuals in dioecious plant populations are often close to unity, or are male biased owing to gender-specific differences in flowering or mortality. Female-biased sex ratios, although infrequent, are often reported in species with heteromorphic sex chromosomes. Two main hypotheses have been proposed to account for female bias: (1) selective fertilization resulting from differential pollen-tube growth of female- versus male-determining microgametophytes (certation); (2) differences in the performance and viability of the sexes after parental investment. Here we investigate these hypotheses in Rumex nivalis (Polygonaceae), a European alpine herb with female-biased sex ratios in which females possess XX, and males XY1Y2, sex chromosomes. Using field surveys and a glasshouse experiment we investigated the relation between sex ratios and life-history stage in 18 populations from contrasting elevations and snowbed microsites and used a male-specific SCAR-marker to determine the sex of nonflowering individuals. Female bias among flowering individuals was one of the highest reported for populations of a dioecious species (mean female frequency = 0.87), but males increased in frequency at higher elevations and in the center of snowbeds. Female bias was also evident in nonflowering individuals (mean 0.78) and in seeds from open-pollinated flowers (mean 0.59). The female bias in seeds was weakly associated with the frequency of male flowering individuals in populations in the direction predicted when certation occurs. Under glasshouse conditions, females outperformed males at several life-history stages, although male seeds were heavier than female seeds. Poor performance of Y1Y2 gametophytes and male sporophytes in R. nivalis may be a consequence of the accumulation of deleterious mutations on Y-sex chromosomes.     

Arbuscular mycorrhiza and water and nutrient supply differently impact seedling performance of dry woodland species with different acquisition strategies

Background: We recently demonstrated that dwarf male plants are much more common among mosses than hitherto thought. Dwarf plants, producing functional male sexual organs occur in 10–20% of the moss species worldwide.

Aims: We investigated how an inadvertent omission of dwarf males affects estimates of male sex expression rates and sex ratios.

Methods: We recorded frequencies of specimens with female-expressing plants, male-expressing normal-sized and/or dwarf plants, and sporophytes in herbarium material of five model species from Macaronesia. We analysed how frequency estimates of male occurrence and sex ratios were affected by excluding dwarf males.

Results: Occurrence of dwarf males was positively associated with sporophyte formation. When dwarf males were excluded, male mate availability was missed in 51–61% of specimens (in 74–76% of sporophyte-containing specimens). Specimen-level sex ratio was balanced in material with sporophytes in the three species exhibiting male dwarfism, but strongly female biased if dwarf males were omitted.

Conclusions: We provide the first quantitative evidence that sex-expressing dwarf males are essential for sexual reproduction and sporophyte production in mosses that form such males. Considering its abundance, male dwarfism therefore deserves more attention among plant biologists. Innovative approaches are required to further study the minute male plants.     

The cost of realized sexual reproduction: assessing patterns of reproductive allocation and sporophyte abortion in a desert moss

The desert moss Syntrichia caninervis exhibits one of the most skewed sex ratios in the plant kingdom, with female individuals far outnumbering male individuals (exceeding 14♀:1♂). The "cost of sex hypothesis" derives from allocational theory and predicts that the sex which is most expensive should be the rarer sex. This hypothesis, which, as considered here represents the realized cost of sexual reproduction, is contingent upon two assumptions that are explored: (1) that male sex expression is more expensive than female sex expression, and (2) that sexual reproduction is resource limited. Using inflorescence biomass and discounting sperm, male sex expression was found to be in the neighborhood of one order of magnitude more expensive than female sex expression, and this difference is reflected in higher numbers of gametangia per male inflorescence, presence of paraphyses in male inflorescences, and a much longer developmental time for male inflorescences. The realized cost of female reproduction from two communities dominated by S. caninervis was found to be lower than the realized cost of male sexual reproduction. Resource-limited reproduction was assessed by determining the frequency of sporophyte abortion, the age distribution of sporophyte abortions, and patterns of sporophyte abortion that may be density dependent. Among ten sexually reproducing populations, abortive sporophytes occurred at a frequency of 0.64. Abortive sporophytes averaged 8% the mass of mature sporophytes, and cohort sporophytes from the same individual female were found to abort in a density-dependent pattern. We conclude that the two assumptions, upon which the cost of sex hypothesis depends, are supported.     

Reproductive biology and population dynamics of wild gametophytes ofEquisetum

This is d study of the reproductive biology of wild gametophytes of Equisetum arvense L., E. fluviatile L., and E. palustre L., which form extensive populations during the summer months on previously inundated bare mud along the margins of reservoirs and lakes in Northern England and Wales. The garnetophytes have all the characteristics of pioneer species: they rapidly attain sexual maturity and are adversely affected by competition. Early in the summer mixtures of male and female gametophytes are produced. The former are smaller and their frequency (which is always less) diminishes throughout the season as a result of bryophytic competition. The rarity of bisexual, initially female, gametophytes, in wild populations is related to the absence of metabolite accumulation which mediates this change in rulture. Significantly different sex ratios between populations and species underlines the labile nature of the sex-determining mechanism in Equisetum. The high frequency of females bearing sporophytes indicates that intergarnetophytic fertilization is highly effective in nature. Absence of any correlation between the incidence of fertilized females and the proportions of males, in conjunction with a consideration of the male gamete dispersal distance, suggests that sporophyte formation is restricted by the availability of ripe archegonia. Sexual reproduction in Equisetum is probably limited by the narrow range of conditions under which gametophytes can become established rather than availability of water for fertilization. The majority of females, which bear one sporophyte, are smaller than unfertilized or polyembryonic females. Their small size results from nutrient demands and allelopathic compounds from the sporophytes which also probably prevent the establishment of gametophytes within mature stands of the parent species. Correlations between female diameters and frequencies of males suggest that gametophytes are more likely to produce archegonia under favourable conditions. The natural reproductive biology of Equisetum is in accord with predictions based on an understanding of the mechanism of sex determination in axenic culture. Several striking parallels between sexuality in Equisetum and dioecious angiosperms are revealed. The absence of winter flooding at two of the gametophyte sites led to the establishment of mature rtands of Equisetum, which produced cones after 3–4 years; two hybrids, E. fluviatile×E. arvense and E. fluviatile×E. palustre, were detected.     

Growth and reproductive characteristics in artificially formed clonal gametophytes ofDryopteris filix-mas (Dryopteridaceae)

Artificially formed clonal gametophytes ofDryopteris filix-mas (L.)Schott were able to grow and reproduce, although growth rates, proportions of hermaphrodites and reproductive efforts were low. Variable density and nutrient levels appeared to affect gametophytic life histories and they continued to influence the viability of the developing sporophyte generation. Differences between populations and sporophytes were discovered in life histories. Hermaphroditic gametophytes were considerably larger and possessed higher viability than did males. Clear differences between clones in the level of phenotypic plasticity in gametophyte size were observed. No neighbor effect on gametophyte sex expression was detected with the densities and nutrient levels used.     

Temporal variations of vegetative features, sex ratios and reproductive phenology in a Dictyota dichotoma (Dictyotales, Phaeophyceae) population of Argentina

This paper addresses the phenology of a Dictyota dichotoma population from the North Patagonian coasts of Argentina. The morphology of the individuals was characterized, and analyses of the temporal variations of vegetative features, diploid and haploid life cycle generations and sex ratios are provided. Individuals, represented by growing sporophytes and gametophytes, occurred simultaneously throughout the year. Morphological variables showed temporal variation, except the width and height of medullary cells, which did not vary between seasons. All vegetative variables were significantly correlated with daylength. Besides, frond length, frond dry mass and apical and basal branching angles were significantly correlated with seawater temperatures. Vegetative thalli were less abundant than haploid and diploid thalli. Sporophytes were less abundant than male and female gametophytes. Male gametophytes dominated in May, August, October and January, and female gametophytes were more abundant in September, November, December, February and March. The formation of female gametangia showed a significant correlation with daylength, and the highest number of gametangia was registered in spring. In general, the male/female sex ratio varied between 1:2 and 1:1. Apical regions were more fertile than basal regions. Our data about frequency in the formation of reproductive structures and male/female ratios are the first recorded in the Dictyota genus and thus could not be compared with populations from other regions of the world. Significant morphological variation was observed in thalli of both life cycle generations, regarding length and dry mass, number of primary branches and branching basal angle. In general, all variables analyzed varied seasonally except cortical cell width.     

GAMETOPHYTE ONTOGENY,SEX EXPRESSION,AND GENETIC LOAD AS MEASURES OF POPULATION DIVERGENCE IN BLECHNUM SPICANT

Early gametophyte ontogeny was quantitatively distinct for Olympic Peninsula, Alaskan, and disjunct Idaho populations of the homosporous fern Blechnum spicant (L.) J. Sm. Although variable, gametophyte sex expression was shown to have a genetic component. Statistically different patterns of sex expression characterize each population. The Olympic Peninsula populations were distinct from each other but consistent in having a predominantly unisexual pattern. The disjunct Idaho population was predominantly bisexual at the time when comparable field collected gametophytes bear sporophytes. Preliminary experiments suggest that an antheridogen operates in this species. Increased sowing density favors maleness, and an extract from soil cultures of gametophytes shifts cultures to an exclusively male pattern after a dramatic suppression of growth. Mating experiments revealed that all populations are interfertile, although fertility was highest when the test Idaho population underwent intergametophytic-selfing. The Idaho population evidenced a low level of genetic load consistent with predictions based on its sex expression. Although Olympic Peninsula populations evidenced apparent high genetic load in some experiments, failure to produce abundant sporophytes in other experiments suggested that additional cultural factors operated to reduce sporophyte formation. Moderate density mating experiments produced single sporophytes that were comparable to field collections. Isolated gametophytes underwent polyembryony after a time delay and gametophyte proliferation. Cultural conditions which allow sporophyte formation on isolated gametophytes without this delay or proliferation must be sought before further genetic analysis is undertaken.     

Sexual reproduction in the phyllodioicous bryophyte Homalothecium lutescens (Hedw.) H.Rob. in relation to habitat age,growth conditions and genetic variation

Sporophyte production and female fertility were investigated in seventeen calcareous grassland demes of the moss Homalothecium lutescens (Hedw.) H.Rob. on the Baltic island of Öland, with the aim of understanding the relationships between sexual reproduction, habitat age, genetic variation and factors related to growth conditions. The overall proportion of fertile female shoots (with perichaetia) was 35%. Fertility status at the level of individual shoots was positively associated with shoot length and density, while within deme fertility was positively associated with bush cover. There was no association between female fertility and habitat age, genetic diversity (H_S) or allelic richness. Out of 1344 investigated shoots, only two were normal-sized fertile males. Dwarf males were also extremely rare, and found almost exclusively on shoots with sporophytes. Few sporophytes were observed (in the two demes with highest fertility and bush cover). No relationship between genetic variation and the frequency of sporophytes and males was found. The lack of a relationship between sexual reproduction and genetic variation suggests that sexual reproduction may not occur in the same grassland fragments as the recruitment of new clones (from spores or vegetative fragments). The majority of the dry, open grassland habitats, where H. lutescens is typically found in the study area, appear to be suboptimal for both dwarf males and fertilization. Sexual reproduction is more likely to occur in shaded (although grazed) grassland patches, where moisture levels are likely to be higher and the moss colonies are generally more vigorous.     

The effect of interspecific mating on sex ratios in the twospotted spider mite and the Banks grass mite (Acarina: Tetranychidae)

Mixed populations of the twospotted spider mite (TSM),Tetranychus urticae (Koch), and the Banks grass mite (BGM),Oligonychus pratensis (Banks), occur on corn and sorghum plants in late summer in the Great Plains. Interspecific matings between these arrhenotokous species occur readily in the laboratory but yield no female offspring. The effect of interspecific mating on female: male sex ratios was measured by examining the F₁ progeny of females that mated with both heterospecific and conspecific males in no-choice situations. TSM females that mated first with BGM males and then with TSM males produced a smaller percentage of female offspring than TSM females that mated only with TSM males (43.1±5.8 and 78.9±2.8% females, respectively). Similarly, BGM females mated with heterospecific males and then with conspecific males produced fewer female offspring than females mated only with BGM males (55.7±5.2 and 77.5±2.5%, respectively). Lower female: male sex ratios were produced also by BGM females that mated with TSM males after first mating with conspecifics (62.4±3.4%). In mixed populations containing males of both species, females also produced lower female: male sex ratios, but these ratios were not as low as expected based on mating propensities and progeny sex ratios observed in no-choice tests. These data suggest that interspecific mating may substantially reduce female fitness in both mite species by reducing the output of female offspring, but in mixed populations this effect is mitigated by unidentified behavioral mechanisms.     

Localization of a Female-Specific Marker on the Chromosomes of the Brown Seaweed Saccharina japonica Using Fluorescence In Situ Hybridization

Background

There is a heteromorphic alternative life in the brown seaweed, Saccharina japonica (Aresch.) C. E. Lane, C. Mayes et G. W. Saunders ( = Laminaria japonica Aresch.), with macroscopic monoecious sporophytes and microscopic diecious gametophytes. Female gametophytes are genetically different from males. It is very difficult to identify the parent of a sporophyte using only routine cytological techniques due to homomorphic chromosomes. A sex-specific marker is one of the best ways to make this determination.

Methodology/Principal Findings

To obtain clear images, chromosome preparation was improved using maceration enzymes and fluorochrome 4′, 6-diamidino-2-phenylindole (DAPI). The chromosome number of both male and female haploid gametophytes was 31, and there were 62 chromosomes in diploid sporophytes. Although the female chromosomes ranged from 0.77 µm to 2.61 µm in size and were larger than the corresponding ones in the males (from 0.57 µm to 2.16 µm), there was not a very large X chromosome in the females. Based on the known female-related FRML-494 marker, co-electrophoresis and Southern blot profiles demonstrated that it was inheritable and specific to female gametophytes. Using modified fluorescence in situ hybridization (FISH), this marker could be localized on one unique chromosome of the female gametophytes as well as the sporophytes, whereas no hybridization signal was detected in the male gametophytes.

Conclusions/Significance

Our data suggest that this marker was a female chromosome-specific DNA sequence. This is the first report of molecular marker localization on algal chromosomes. This research provides evidence for the benefit of using FISH for identifying molecular markers for sex identification, isolation of specific genes linked to this marker in the females, and sex determination of S. japonica gametophytes in the future.     

Life history variation in gametophyte populations of the moss Ceratodon purpureus (Ditrichaceae)

The life cycles of mosses and other bryophytes are unique among land plants in that the haploid gametophyte stage is free-living and the diploid sporophyte stage is ephemeral and completes its development attached to the maternal gametophyte. Despite predictions that populations of haploids might contain low levels of genetic variation, moss populations are characterized by substantial variation at isozyme loci. The extent to which this is indicative of ecologically important life history variation is, however, largely unknown. Gametophyte plants from two populations of the moss Ceratodon purpureus were grown from single-spore isolates in order to assess variation in growth rates, biomass accumulation, and reproductive output. The data were analyzed using a nested analysis of variance, with haploid sib families (gametophytes derived from the same sporophyte) nested within populations. High levels of life history variation were observed within both populations, and the populations differed significantly in both growth and reproductive characteristics. Overall gametophytic sex ratios did not depart significantly from 1:1 within either population, but there was significant variation among families in both populations for progeny sex ratio. Some families produced predominantly male gametophytes, while others yielded predominantly females. Because C. purpureus has a chromosomal mechanism of sex determination, these observations suggest differential (but unpredictable) germination of male and female spores. Life history observations showed that male and female gametophytes are dimorphic in size, maturation rates, and reproductive output.     

Genetic diversity,sexual condition,and microhabitat preference determine mating patterns in Sphagnum (Sphagnaceae) peat‐mosses

In bryophytes, the possibility of intragametophytic selfing creates complex mating patterns that are not possible in seed plants, although relatively little is known about patterns of inbreeding in natural populations. In the peat‐moss genus Sphagnum, taxa are generally bisexual (gametophytes produce both sperm and egg) or unisexual (gametes produced by separate male and female plants). We sampled populations of 14 species, aiming to assess inbreeding variation and inbreeding depression in sporophytes, and to evaluate correlations between sexual expression, mating systems, and microhabitat preferences. We sampled maternal gametophytes and their attached sporophytes at 12–19 microsatellite loci. Bisexual species exhibited higher levels of inbreeding than unisexual species but did generally engage in some outcrossing. Inbreeding depression did not appear to be common in either unisexual or bisexual species. Genetic diversity was higher in populations of unisexual species compared to populations of bisexual species. We found a significant association between species microhabitat preference and population genetic diversity: species preferring hummocks (high above water table) had populations with lower diversity than species inhabiting hollows (at the water table). We also found a significant interaction between sexual condition, microhabitat preference, and inbreeding coefficients, suggesting a vital role for species ecology in determining mating patterns in Sphagnum populations. © 2015 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 115 , 96–113.     

A SCAR MOLECULAR MARKER SPECIFICALLY RELATED TO THE FEMALE GAMETOPHYTES OF SACCHARINA (LAMINARIA) JAPONICA (PHAEOPHYTA)1

PCR amplification was employed to identify female or male gametophyte associated markers in Saccharina japonica (Aresch.) C. E. Lane, C. Mayes et G. W. Saunders (=Laminaria japonica Aresch.). One pair of the primers, P5, was screened from five pairs designed based on a specific sequence (GenBank accession no. AB069714 ) of Marchantia polymorpha Y chromosome, resulting in a differential band ～500 bp in size between female and male gametophytes of Rongfu strain of S. japonica. According to the SCAR (sequence‐characterized amplified regions) strategies, one pair of primers, P51, was designed on the basis of the sequence of this band that was only present in female gametophytes. A SCAR marker, designated FRML‐494 (494‐bp Female‐Related Marker of S. japonica, GenBank accession no. EU931619 ), was developed successfully by PCR amplification using the designed P51 primer pair. The SCAR marker was verified to be present only in female gametophytes of another variety 901 of this kelp that was a hybrid between S. japonica as paternal and S. longissima (Miyabe) C. E. Lane, C. Mayes, Druehl et G. W. Saunders (=Laminaria longissima Miyabe) as maternal, suggesting that the FRML‐494 marker was specifically related to female gametophytes of the genus. This marker is the first molecular tool reported for sex identification in kelps. This study was beneficial for identifying gametophyte gender during vegetative growth and for judging whether the monogenetic sporophytes came from exclusive male or female gametophytes, as well as for further research on sex determination at the molecular level in kelps.     

葫芦藓（Funaria hygrometrica）配子体和孢子体重金属富集特性比较

通过对湘西茶田钒矿废弃冶炼厂矿渣上葫芦藓的野外生态调查和采集,利用原子吸收光谱仪、电感耦合等离子发射光谱仪和原子荧光光谱仪分析了葫芦藓配子体和孢子体及其基质重金属含量。结果表明葫芦藓配子体和孢子体富集了大量的重金属,各重金属元素在配子体和孢子体间的富集存在较大的差异,配子体比孢子体显著富集重金属元素（p<0.05）,Zn和Mn在葫芦藓植物体中比其他重金属元素更高。同时也讨论了重金属在苔藓植物中的富集及生物阻抗的作用。     

NORTHERN AND SOUTHERN HEMISPHERE HYBRIDS OF MACROCYSTIS (PHAEOPHYCEAE)1

All combinations of individuals of Macrocystis pyrifera (L.) C. Agardh, M. integrifolia Bory, and M. angustifolia Bory hybridized. Gametophyte isolates obtained from 18 individuals were used, including M. pyrifera and M. integrifolia from the extremes of their Northern Hemisphere ranges along the Pacific Coast of North America and M. pyrifera and M. angustifolia from Tasmania, Australia. All combinations of gametophytes produced sporophytes of normal morphology, with the exception of crosses involving three gametophyte isolates. One female (M. integrifolia) and two male (M. pyrifera and M. angustifolia)gametophyte isolates were unable to produce normal sporophytes in combination with gametophytes of the opposite sex. Some cultures of female gametophytes produced abnormally shaped parthenogenetic sporophytes. Gametophytes and sporangia of M. pyrifera had n= 16 chromosomes. The M. integrifolia female gametophyte that was unable to produce normal sporophytes had n = ca. 32 chromosomes. These results show that these species of Macrocystis have not become reproductively isolated. Although these species may be considered conspecific according to the biological species concept, we recommend that they continue to be recognized as separate species based on morphological differences.     

PROBLEMS IN THE IDENTITY AND ORIGIN OF THE APPALACHIAN VITTARIA GAMETOPHYTE,A SPOROPHYTELESS FERN OF THE EASTERN UNITED STATES

Populations of a fern gametophyte presumed to be of the genus Vittaria occur commonly in the uplands of the southeastern United States. The gametophytes occur on non-calcareous rock outcrops of various composition in areas which provide continuous moisture and protection from temperature extremes. Gametophytes in these habitats are robust and long lived, frequently forming the dominant vegetation in areas covering several square feet. Reproduction is exclusively vegetative by production of gemmae. Although sex organs are present in most populations, viable sporophytes are never produced. The gametophytes have previously been considered most likely to be V. lineata, which occurs in Florida, but morphological and physiological comparisons do not support that conclusion. Significant differences between Appalachian and Florida gametophytes occur in growth form, growth rate, cold hardiness, sporophyte production, and patterns of gemma production. The distribution of the Appalachian gametophytes correlates with old, unglaciated land masses suggesting antiquity rather than recent introduction. Present evidence favors the interpretation that the Appalachian Vittaria gametophytes either belong to a tropical American species from which they have long been separated, or that they represent a distinct species of which the sporophyte no longer exists.     

Growth and reproductive responses of Laminaria longicruris (Laminariales,Phaeophyta) to nutrient enrichment

A series of comparative culture experiments were conducted in order to determine responses of Laminaria longicruris male and female gametophytes and juvenile sporophytes to several temperatures (5, 10, 15, 20 °C), light levels (10, 35, 75 µmol m^–2 s^–1) and media nitrogen concentrations (0, 20, 100 µM ammonium-nitrogen). Responses were measured as numbers of male and female gametophytes producing gametangia and number of sporophytes produced following fertilization. Both male and female gametogenesis was reduced at 5 and 20 °C versus 10 and 15 °C. At 20 °C gametogenesis inhibition was greater with higher levels of ammonium-nitrogen concentration (100 µM). Sporophyte production was more sensitive to light, temperature and nitrogen concentration than gametogenesis. Production of sporophytes was inhibited completely at 20 °C. At lower temperatures, increasingly higher nutrient concentrations produced greater inhibition of production of sporophytes.     

Sex‐linked microsatellite marker detected in the female gametophytes of Undaria pinnatifida (Phaeophyta)

In our microsatellite analysis of three male and three female gametophytes of Undaria pinnatifida (Harv.) Suringar, a microsatellite marker (part of the locus Up‐AC‐2A8, GenBank accession no. AY738602.1) was only polymerase chain reaction‐amplified in three female gametophytes. This putative female‐specific marker was further tested by the use of 32 male and 21 female gametophytes maintained in the Marine Biological Culture Collection Centre, China. In addition, three sporophytes were included for confirmation. Results showed that the marker was present in all of the female gametophytes and sporophyte cultures, but absent in all of the male gametophytes. To our knowledge, this is the first sex‐related marker ever reported in U. pinnatifida. The discovery of this marker will accelerate gender identification and shed light on our understanding of the mechanisms of sex determination at a molecular level in this commercially important seaweed.     

Expression of the<Emphasis Type="Italic"> lacZ</Emphasis> reporter gene in sporophytes of the seaweed<Emphasis Type="Italic"> Laminaria japonica</Emphasis> (Phaeophyceae) by gametophyte-targeted transformation

The seaweed Laminaria japonica (Phaeophyceae) has a two-generation life cycle consisting of haploid gametophytes and diploid sporophytes. Female and/or male gametophytes were transformed using particle bombardment and the histological LacZ assay was performed on sporophytes generated by either parthenogenesis or inbreeding. Female gametophyte-targeted transformation resulted in similar lower efficiencies in both parthenogenetic and zygotic sporophytes, and only a chimeric expression pattern was observed. Male gametophyte-targeted transformation led to a higher efficiency, with 3.5% of the zygotic sporophytes stained completely blue (all-blue), implying the integration of lacZ at the one-cell stage. Polymerase chain reaction analysis using primers specific for a lacZ-vector juncture fragment and subsequent blotting indicated the presence of the introduced gene in the sporophytes. The method reported here has a potential for seaweed transformation using spore-based bombardment followed by the developmental process.Abbreviations DPR Detected positive rate - ER Expression rateCommunicated by F. Sato