In Vitro Culture of Orobanche aegyptiaca

Calli derived from seedlings of the parasitic plant Orobancheaegyptiaca were grown on enriched nutrient agar. Growth wasenhanced by the addition of 0.1 mg 1^–1 NAA and 5 % coconutmilk. The calli developed two kinds of elongated protrusions:thin ones which resemble the young root radicle and thick oneswhich resemble the adventitious roots of Orobanche plants. Bothformed pad-like structures when touching the substrate. In liquidculture, some of the calli were observed to surround isolatedcultured tomato roots. Orobanche aegyptiaca Pers., in vitro culture, germination, coconut milk, root parasite     

The Effects of Root Pruning on the Water Relations of Helianthus annuus L.

Four-week-old Helianthus annuus plants, grown in both soil andliquid culture, were root pruned at the point of root attachmentto the stem. Transpiration, leaf water potential and leaf conductivitywere monitored for several days after pruning. Pruning loweredtranspiration and leaf conductivity in amounts proportionalto the amount of pruning. In some experiments pruning causeda slight lowering of leaf water potential, while in others nopruning effect could be found. The effects of pruning varieddepending upon culture conditions, with greater effects beingfound in soil and unaerated liquid culture than in aerated liquidculture. Soil water potential did not appear to have a stronginfluence on the magnitude of the pruning effect. The effectsof root pruning are less than would be predicted by an Ohm'sLaw analysis of flow; possible reasons for this are discussed.     

On separation and identity of fern antheridiogens

Chromatographic studies show that the hormones controlling antheridiuminduction in the fern species Pteridium aquilinum (Polypodiaceae),Anemia phyllitidis and Lygodium japonicum (Schizaeaceae) aredifferent molecular entities. SCHRAUDOLF's report that gibberellic acid induces antheridiain A. phyllitidis and L. japonicum was confirmed. The activityspectrum of GAs towards species of different fern families stronglyresembles that of the native Anemia antheridiogen. However,the native antheridiogens of A. phyllitidis, and of Lygodiumjaponicum, are more species-selective in their action than isGA₃. Preliminary studies have yielded no conclusive evidenceon whether the native antheridiogens are gibberellins. (Received August 21, 1967; )     

Suppression of Inhibitory Effects of Copper on Enzymatic Activities by Copper-Binding Substances from Athyrium yokoscense Assayed in vitro

A metal-tolerant fern, Athyrium yokoscense, is capable of growingin highly copper-contaminated soil, but cupric chloride inhibitedthe activities of some enzymes extracted from the fern. Thefunction in the detoxification of copper of two copper-bindingsubstances was investigated by examination of their effectson various enzymes assayed in vitro, i.e. acid phosphatase (orthophosphoric-monoesterphosphohydrolase [acid optimum], EC 3.1.3.2 [EC] ), glucose-6-phosphatedehydrogenase (D-glucose-6-phosphate: NADP⁺ 1-oxidoreductase,EC 1.1.1.49 [EC] ) and isocitrate dehydrogenase (threo-Ds-isocitrate:NADP⁺ oxidoreductase [decarboxylating], EC 1.1.1.42 [EC] ). The twocopper-binding substances, whose apparent molecular weightsare 9.5 kDa and 2 kDa, were previously obtained from the solublecytoplasmic fraction of the fern root. The 9.5-kDa substance,which is a cysteine-rich peptide induced as a result of exposureof the fern to copper, was found to suppress almost entirelythe inhibitory effects of the metal on the enzymes. The suppressoractivity of the peptide was nearly as effective as that of ethylenediaminetetraaceticacid. The 2-kDa substance, which is also found in fern thathas not been exposed to copper, had a more modest suppressoractivity. These results indicate that the 9.5-kDa substancemay contribute to the copper-tolerance of the fern growing incopper-contaminated soil. (Received August 26, 1988; Accepted March 17, 1989)     

An Inducible Copper-Thiolate Complex in the Fern, Athyrium yokoscense: Involvement in Copper-Tolerance of the Fern

Two peaks of Cu-containing material were isolated from the solublecytoplasmic fraction of roots of Athyrium yokoscense, a fernwith a tolerance for heavy metals, by gel filtration on Bio-GelP-30 and Sephadex G-25. Peak I (apparent molecular weight 9.5kDa) was only observed in the case of ferns growing on Cu-contaminatedsoil. Peak II (apparent molecular weight 2 kDa) was found inextracts of ferns from both Cu-contaminated and uncontaminatedsoil. Peak I was induced in the fern collected from an uncontaminatedarea by cultivation in Cu-contaminated soil for 14 months. Theinduced Cu-binding complex present in peak I, therefore, maybe involved in mechanisms of Cu-tolerance in the fern. The complexwas purified by column chromatography on DEAE-cellulofine andsubsequent high-performance liquid chromatography. The purifiedcomplex contained a high percentage (26.8%) of cysteine, typicalof metal-binding peptides from plants. (Received June 1, 1988; Accepted September 12, 1988)     

Studies on the nutrition of rice cell culture I. A simple, defined medium for rapid growth in suspension culture

We investigated the nutrient requirements of rice in liquidculture and developed a revised medium of mineral salts, sucrose,thiamine and 2,4-dichlorophenoxyacetic acid. The following majornutrients at the indicated concentrations were beneficial: NO₃-N(40 mM), NH₄-N (5.0 mM), P (2.0 mM) and K (40 mM). Cobalt, iodine,pyridoxine, nicotinic acid and m-inositol were not essentialand were excluded from the revised R-2 medium. Growth was betterwith ammonium and nitrate together than with nitrate as thesole nitrogen source. Cell growth in the R-2 medium was superiorto that obtained in B5, Heller, Murashige-Skoog and White media. (Received March 31, 1973; )     

The Spontaneous Formation of Antheridia in Onoclea is not Blocked by Light

Onoclea sensibilis gametophytes were grown from spores on ashedsoil and agar to determine if the spontaneous formation of antheridiacan be blocked by light. Under most conditions, dark-grown gametophytesformed antheridia later than or at the same time as gametophytesgrown in the light. Under no circumstances was there a rapidonset of maleness in the dark. These results contradict thehypothesis that, in Onoclea, antheridiogen is required to inducemaleness because light inhibits the formation of antheridia.In the light, antheridia formed on heart-shaped thalli. In darkness,antheridia formed on filamentous gametophytes. The timing ofonset of maleness was affected by temperature and the presenceof sucrose. The effect of sucrose on the comparison betweenlight and dark treatments depended on both substrate and temperature Onoclea sensibilis, L., sensitive fern, fern gametophytes, sexuality, light-induced block     

Phytochrome Regulation of Expression of mRNA Encoding the Major Light-Harvesting Chlorophyll a/b-Binding Proteins of Photosystem II in the Haploid Phase of Adiantum capillus-veneris

Light regulates development in the fern Adiantum capillus-venerisduring both the haploid gametophyte and diploid sporophyte phases.In this study, we have investigated the role of the photoreceptorphytochrome in haploid spore germination and regulation of expressionof mRNA encoding the major light-harvesting polypeptides ofphotosystem II {Lhcb mRNAs). Both of these responses appearto be induced by phytochrome and inhibited by blue light, acting.throughseparate photoreceptor(s). Characteristics of phytochrome actionwere similar for both responses, showing no induction in thevery low fluence range and an ‘escape’ from far-redreversal for 50% of the response by 6 h. Overall, these resultsshow that phytochrome regulates both a morphological and a molecularprocess in haploid fern spores, and that these processes mayshare a common signal transduction pathway. (Received February 4, 1998; Accepted April 8, 1998)     

Gametophytic and Sporophytic Regeneration from Bud Scales of the FernPlatycerium bifurcatum(Cav.) C.Chr.In Vitro

Regeneration fromPlatycerium bifurcatumscalesin vitrowas observedfor the first time. Regeneration was obtained from scales detachedfromin vitrogrown juvenile shoots. Regeneration began as rhizoidsor as a proliferation of one or more scale cells into undifferentiatedoutgrowths distinguishable from adjacent cells by their shapeand colour. Outgrowths developed into adventitious buds, aposporousgametophytes, or remained undifferentiated. Numerous outgrowthsformed rhizoids on their surfaces. Aposporous gametophytes producedgametangia. Sucrose enhanced scale viability and regeneration,and promoted rhizoid development on outgrowths, but sucroseconcentrations greater than 0.1% tended to inhibit apospory. Platycerium bifurcatum(Cav.) C.Chr.; staghorn fern; scale; regeneration; organogenesis; sucrose; apospory     

Developmental Morphology of Platycerium coronarium (Koenig) Desv

The growth habit and certain developmental aspects of Platyceriumcoronarium (stag's horn fern) are described. Starting from theprimordial stage, the nest and pendulous leaves develop to maturityin 90 and 80 days respectively. The fertile lobe, which is partof the pendulous leaf, reaches its maximum size in 40 days.The morphogenesis of the nest leaf is more variable, and itmatures and deteriorates earlier than the pendulous leaf. Theacrostichoid sorus formation is completed in 3 weeks from inceptionand spore dispersal takes place when the fertile lobe is about100 days old. The area of the fertile lobe and number of sporesproduced were determined. On Knop's agar medium the gametophytesdevelop in 2 months and 85 per cent of them are unisexual (bothmale and female) and 15 per cent bisexual. Less than 1 per centof the gametophytes give rise to sporophytes. The juvenile leavesare simple, displaying open dichotomous venation; the firstnest and pendulous leaves are produced 24 months after the dateof spore germination. Platycerium coronarium, stag's horn fern, leaf development, morphogenesis, spore production     

Photomorphogenesis in Pteris vittata IV. Action spectra for inhibition of phytochrome-dependent spore germination

Action spectra between 350 and 500 nm for the inhibition ofphytochrome-dependent spore germination in the fern Pteris vittatawere obtained. Both action spectra obtained before and afterred light irradiation have peaks at about 440 nm and 380 nmand shoulders from 440 to 480 nm. These results suggest thatthe phytochrome system is not involved in the inhibitory processof spore germination induced by short irradiation with bluelight. (Received October 8, 1970; )     

Dispersed Fern Sporangia from the British Tertiary

Dispersed fern sporangia have been recovered from the BembridgeMarls of the Isle of Wight and from Bracklesham deposits ina borehole West of Poole, Dorset. These fern sporangia confirmthe presence of the family Polypodiaceae s.1. in the BritishTertiary flora. Previously records of this family have beenbased solely on vegetative material, usually fragmentary, forwhich taxonomic assignment is inevitably insecure. The sporangia,containing spores and with adherent paraphyses, are shown tobe most comparable with the Recent genus Acrostichum L. andare described as Acrostichum anglicumsp. nov. The distributionand occurrence of these sporangia suggests the extensive developmentof Acrostichum in the lacustrine environments of Bembridge times.     

Gene Activity during Germination of Spores of the Fern, Onoclea sensibilis: RNA and Protein Synthesis and the Role of Stored mRNA

Pattern of ³H-uridine incorporation into RNA of spores of Onocleasensibilis imbibed in complete darkness (non-germinating conditions)and induced to germinate in red light was followed by oligo-dTcellulose chromatography, gel electrophoresis coupled with fluorographyand autoradiography. In dark-imbibed spores, RNA synthesis wasinitiated about 24 h after sowing, with most of the label accumulatingin the high mol. wt. poly(A)^–RNA fraction. There was noincorporation of the label into poly(A) ⁺ RNA until 48 h aftersowing. In contrast, photo-induced spores began to synthesizeall fractions of RNA within 12 h after sowing and by 24 h, incorporationof ³H-uridine into RNA of irradiated spores was nearly 70-foldhigher than that into dark-imbibed spores. Protein synthesis,as monitored by ³H-arginine incorporation into the acid-insolublefraction and by autoradiography, was initiated in spores within1–2 h after sowing under both conditions. Autoradiographicexperiments also showed that the onset of protein synthesisin the cytoplasm of the germinating spore is independent ofthe transport of newly synthesized nuclear RNA. One-dimensionalsodium dodecyl sulphate-polyacrylamide gel electrophoresis of³⁵S-methionine-labelled proteins revealed a good correspondencebetween proteins synthesized in a cell-free translation systemdirected by poly(A) ⁺RNA of dormant spores and those synthesizedin vivo by dark-imbibed and photo-induced spores. These resultsindicate that stored mRNAs of O. sensibilis spores are functionallycompetent and provide templates for the synthesis of proteinsduring dark-imbibition and germination. Key words: Onoclea sensibilis, fern spore germination, gene expression, protein synthesis, sensitive fern, stored mRNA     

Characterization of Protein Synthetic Changes in a Desiccation-Tolerant Fern, Polypodium virginianum. Comparison of the Effects of Drying, Rehydration and Abscisic Acid

The desiccation-tolerant fern, Polypodium virginianum, was ableto lose more than 60% of its fresh weight during periods ofprolonged water stress and fully recover during periods of wateravailability. Rehydration from the air-dry state occurred rapidly(within 24 h) with a concurrent initiation of protein synthesis.A low-molecular-weight doublet was synthesized during waterstress and was stable for at least 24 h after rehydration. Theseproteins were expressed also when the fully hydrated frondswere subjected to exogenous ABA even though drying resultedin a 10-fold reduction in ABA content. The large subunit ofRubisco was synthesized during drying, but despite temporaryaccumulation, the de novo synthesized component was degradedduring rehydration. During rehydration from the air-dry state,unique rehydration-specific polypeptides were produced. Thesepolypeptides were synthesized at different water contents andtimes of rehydration, and were transiently expressed. This transientexpression may mean that they are only necessary during theearly stages of rehydration when the rapid initiation of physiologicaland repair processes is essential. The response of this fernto desiccation has features which are similar to those reportedfor desiccation-tolerant mosses, and others which are reminiscentof desiccation-tolerant angiosperms. Key words: Desiccation-tolerance, fern, Polypodium virginianum, protein synthesis, abscisic acid     

Isolated Megaspore Tetrads of Stauropteris burntislandica

A new type of fossil spore tetrad, Didymosporites scotti gen.et sp. nov. is described from the Dinantian and Namurian (Carboniferous)of Ireland, Scotland, and England. The spores always occur inthe form of a tetrad composed of two large (presumably fertile)and two minute (presumably abortive) spores, all with cutinizedwalls. Reasons are given for regarding these as the megasporesof the coenopterid fern Stauropteris burntislandica; their distributionindicates a far wider range in time and space than that previouslyknown for this species. Their occurrence as adhering tetradssuggests that Stauropteris burntislandica 'shed' its megasporesstill enclosed in the sporangiumIan interesting analogy withboth Lepidocarpon and the seeds of Pteridosperms.     

Effects of Sodium Selenite on the Growth of Bracken Fern Root Callus

Sodium selenite promotes the growth of bracken fern root callusgrowth over a narrow, low range of concentrations, while higherconcentrations are strongly inhibitory. Pteridium aquilinum, bracken fern, sodium selenite, selenium, callus     

Changes in the Level of the Protein Nitrogen during Growth of the Gametophyte and the Initiation of the Sporophyte of Dryopteris borreri Newm.

Estimations of the amounts of organic nitrogen in spores anddeveloping gametophytes of Dryopteris borreri, a fern reproducingapogamously and lacking archegonia, have shown that the nitrogenin the gametophytes increases exponentially. The logarithmicrate of increase of the nitrogen remains unchanged during theinitiation and emergence of the sporophyte. Gametophytes aboutto produce sporophytes yielded only filamentous growths whentransferred to a medium containing 8-azaguanine, and the increasein their content of organic nitrogen was very small. The resultsare discussed in relation to similar estimations made by otherworkers upon the gametophytes of Dryopteris erythosora(Eat.)Schott, a fern reproducing sexually. The interpretation placedupon the results obtained for D. erytkrosora is questioned,and a new interpretation of these results, together with thosefor D. borreri, is related to the difference in morphology ofthe two generations in the life cycle of the Polypodiaceousferns.     

Control of the Entry into S Phase by Phytochrome and Blue Light Receptor in the First Cell Cycle of Fern Spores

Phytochrome- and a blue light receptor-dependent pathway antagonisticallyregulate the first mitosis in spores of the fern Adiantum capillus-venerisL. This study focused on determining which phase(s) of the cellcycle is positively regulated by phytochrome and negativelyregulated by a blue light receptor in germinating spores. Incorporationof the radioactivity of ³H-thymidine into the acid-insolublematerial prepared from the spores indicated that phytochromein the PFR form induced the entry into S phase of the firstcell cycle in the spores 20-28 h after irradiation with redlight. Blue light treatment before or after red light treatmenttotally prevented the P_FR-induced DNA synthesis. Brief irradiationwith red, far-red or blue light showed no effects on mitosisif the irradiation was given 28 h after the red light induction,during S and M phases. These results indicate that phytochromeand a blue light receptor regulate the entry into S phase duringthe first cell cycle of fern spores. ( Accepted July 10, 1997)     

Control of heat-shock response of young gametophytes of the sensitive fern is at the translational level

Young gametophytes of the sensitive fern, Onoclea sensibilis,respond to heat-shock by synthesizing in excess certain proteinsthat are made at normal growth temperature. Enhanced proteinsynthesis occurred during a 2 h heat-shock at a range of temperaturesbetween 38 °C and 50 °C. Although a temperature of 50°C proved lethal, a 5 min pulse at 50 °C resulted inenhanced synthesis of heat-shock proteins which continued forseveral hours at 25 °C. After heat-shock at 50 °C for10 or 15 min, the gametophytes temporarily lost their capacityfor protein synthesis but normal protein synthesis was resumedwithin 24 h of heat-shock. A heat-shock at 38 °C precedingone at 50 °C did not have any protecting effect on the gametophytes.In vitro translation of poly(A)+ RNA isolated from heat-shockedgametophytes yielded several proteins including heat-shock proteins.The results suggest that, rather than activating genes encodingnew messages for the synthesis of stress proteins, heat-shockof gametophytes of O. sensibilis triggers a controlling systemwhich enhances the translation of certain messages that aresynthesized at normal growth temperature. Key words: Onoclea sensibilis, heat-shock response, protein synthesis, sensitive fern, in vitro translation     

Effects of Population Density on Sex Expression in Onoclea sensibilis L. on Agar and Ashed Soil

Population density affected the sex expression of agar-growngametophytes of Onoclea sensibilis L. The time of onset of sexualitywas advanced, the proportion of females was increased, and thegrowth rate of individuals was greater at lower densities. Populationdensity had no effect on the sex expression of Onoclea grownon ashed soil, and there was no difference in growth rate ofindividuals grown on ashed soil at different densities. Covariateanalysis, using thallus width as a measure of growth rate, indicatedthat the effect of density on sex expression was mostly associatedwith growth rate. The differing effects of population densityon agar and ashed soil demonstrate that substrate influencessex expression in Onoclea. This influence is most dramatic insingle-gametophyte cultures, where agar cultures produced 97per cent females and ashed soil cultures 100 per cent males. Onoclea sensibilis L., sensitive fern, fern gametophytes, sexuality, population density