荷叶铁线蕨孢子的不同贮藏时间与培养条件对配子体和孢子体形成的影响

研究了荷叶铁线蕨(Adiantum reniforme var.sinense)的不同贮藏时间与培养条件对其配子体发育和孢子体形成的影响,结果表明:荷叶铁线蕨孢子为不含叶绿素孢子,在室温下保存5年以上时间依然具有活力,但不同的贮藏时间对配子体和孢子体的形成具有显著的影响。其中,室温贮藏7d的配子体形成的时间为29d,而贮藏5年的为38d;贮藏7d的孢子体形成的时间为96d,贮藏4个月的为105d。在培养室[(25±1)℃恒温和每天2000~2500lx12h光照]沙基质中,从孢子播种到形成孢子体时间最短,为87d;沙基质比腐殖土更利于孢子体的形成;配子体密度在不超过3株/cm2的情况下,密度越高越有利于孢子体的形成。     

<Emphasis Type="Italic">In vitro</Emphasis> culture of <Emphasis Type="Italic">Drynaria fortunei</Emphasis>, a fern species source of Chinese medicine “Gu-Sui-Bu”

This study reports spore germination, early gametophyte development and change in the reproductive phase of Drynaria fortunei, a medicinal fern, in response to changes in pH and light spectra. Germination of D. fortunei spores occurred on a wide range of pH from 3.7 to 9.7. The highest germination (63.3%) occurred on ½ strength Murashige and Skoog basal medium supplemented with 2% sucrose at pH 7.7 under white light condition. Among the different light spectra tested, red, far-red, blue, and white light resulted in 71.3, 42.3, 52.7, and 71.0% spore germination, respectively. There were no morphological differences among gametophytes grown under white and blue light. Elongated or filamentous but multiseriate gametophytes developed under red light, whereas under far-red light gametophytes grew as uniseriate filaments consisting of mostly elongated cells. Different light spectra influenced development of antheridia and archegonia in the gametophytes. Gametophytes gave rise to new gametophytes and developed antheridia and archegonia after they were transferred to culture flasks. After these gametophytes were transferred to plastic tray cells with potting mix of tree fern trunk fiber mix (TFTF mix) and peatmoss the highest number of sporophytes was found. Sporophytes grown in pots developed rhizomes.     

Seed abortion in the sexual counterpart of <Emphasis Type="Italic">Brachiaria brizantha</Emphasis> apomicts (Poaceae)

Gametophytic organization, fertilization and reproductive success are described for the fertile diploid Brachiaria brizantha accession BRA-002747 which is being raised for use in Brachiaria breeding programs, as well as to understand and control of apomixis in this genus. The current paper reports on reproductive biology and analysis of seed set in field experiments during three consecutive years. Unsuccessful seed production in this plant is believed to correlate with early inbreeding depression, based on the reproductive features analyzed. Caryopsis development was observed using differential interference contrast microscopy with seed set determined by the number of self- and open-pollinated pistils that fully developed into viable seeds. Developing and mature female and male gametophytes were observed in the context of flower phenology, morphology and anthesis patterns. Pollen viability was determined by acetocarmine staining and by observation of germination in vivo, which was also used to observe pollen tube/pistil interaction. Although normal development was observed in floral structures, anthesis and gametophytes, seed set was low, with 2 and 6% in self- and open-pollination, respectively, producing seed. Variations observed in the female organs, such as the presence of a hermaphrodite flower in 50% of the inferior floscules and the presence of multiple embryo sacs of the Polygonum type within the same ovule in 15% of the pistils, are not related to low fertility. The majority of pollen grains are viable, in spite of the reduced number of pollen tubes within the style and ovary carpel, and a developing caryopsis was observed in 70% of self-pollinated pistils, indicating successful double fertilization from 2 days after anthesis (DAA). Nevertheless, abortion gradually increased from 2 until 7 DAA and remains elevated until 12 DAA, when caryopsis maturity is achieved. These data confirm low seed set in this accession and indicate that low fertility is not a consequence of abnormalities, either in the floral or gamete structures, or pollen tube rejection, but most likely a consequence of inbreeding depression.     

苏铁蕨配子体发育的研究

用无机培养基培养苏铁蕨(Brainea insignis (Hook.)J.Sm.)的孢子,显微镜下观察记录其孢子萌发及配子体形态发育过程.结果表明:孢子褐色,单裂缝,具周壁,稍褶皱.接种3 d左右孢子萌发,萌发类型为书带蕨型(VittariaType),原叶体发育类型为槲蕨型(Drynaria Type).接种15 d左右发育为片状体.接种25 d左右发育为成熟原叶体,呈心形,其翼面和翼缘均分布有毛状体.精子器由3细胞构成,成熟颈卵器颈部由4列细胞组成,4～5层细胞高.     

Photolithotrophic cultivation of <Emphasis Type="Italic">Laminaria japonica</Emphasis> gametophyte cells in a silicone tubular membrane-aerated photobioreactor

Gametophyte cells of brown algae Laminaria japonica were employed both in a modified silicone tubular membrane-aerated photobioreactor (bubble-less cultivation mode) and a bubble-column photobioreactor (bubbling cultivation mode), to study different gas–liquid mixing modes on cell growth rate and cell physiological status. With an inoculum density of 50 mg DCW l⁻¹, in modified artificial Pacific seawater (APSW) medium at 13°C, light intensity of 60 μE m⁻² s⁻¹, light cycle of 16/8 h L/D, and aeration rate of 60 ml min⁻¹, the specific growth rates were 0.082 d⁻¹ for bubble-less mode and 0.070 d⁻¹ for bubbling mode with biomass, in the form of dry cell density, increasing 10.9 and 6.8 times, respectively, during the 36 days’ photolithotrophic cultivation. The specific oxygen evolution rate under bubble-less mode was 39.6% higher than under bubbling mode on the 18th day. The gametophyte cells grew in cell aggregates with clump sizes, at day 36, of 1.5 mm and 0.5 mm diameter under bubble-less and bubbling mode respectively and cell injury percentages of 5.1% and 21.1%, respectively. The silicone tubular membrane-aerated photobioreactor was better suited for the cultivation of fragile macroalgal gametophyte cells due to the absence of hydrodynamic shear stress caused by fluid turbulence and the presence of a bubble-less gas supply.     

The effect of cytokinins on growth and sexual organ development in the gametophyte of <Emphasis Type="Italic">Blechnum spicant</Emphasis> L.

In this study, we report the role of exogenous and endogenous cytokinins on growth and sexual organ development in the fern Blechnum spicant L. Spore-derived gametophytes (SG) were cultured in full-strength Murashige and Skoog (1962) liquid medium supplemented with (a) 4.44 μMN⁶-benzyladenine (BAP), (b) a crude extract from mature female gametophytes, and (c) 4.44 μM BAP in combination with the crude extract from mature gametophytes, respectively. Both BAP and the crude extract delayed the gametophyte development, and this effect was increased when they were added together. With respect to sexual organ development, BAP inhibited the sexual organ formation, while the crude extract favored antheridia formation; however, when added together, the percentage of antheridia decreased. The endogenous level of the cytokinins cis-zeatin (cZ), cis-zeatin-riboside (cZR), dihydrozeatin (DHZ), dihydrozeatin riboside (DHZR), isopentenyl adenine (iP), isopentenyl adenosine (iPR), isopentenyl-9-glucoside (iP9G), trans-zeatin (tZ), and trans-zeatin riboside (tZR) were analyzed in female and male gametophytes of B. spicant L. The endogenous levels of cytokinins tZ, cZ, DHZ, cZR, iP, and iPR were higher in female gametophytes than in male gametophytes, with the endogenous iP and iPR content being increased more than 300 and 400 times, respectively.     

Germination and reproductive biology in salty conditions of Asplenium marinum (Aspleniaceae), a European coastal fern

Although there are several species of ferns that are important components of many coastal ecosystems, the adaptations that allow these species to live in salty conditions have been studied only in two Acrostichum species. We have studied the effects of salt on germination, development of the gametophyte and sexual expression of the fern Asplenium marinum, which lives on coastal cliffs of Europe. Cultures at four different levels of salt concentration, 0%, 0.98%, 1.96% and 3.26%, were established for this purpose. Salt caused a significant delay in spore germination, and prevented it completely at high concentrations (3.26%). However, spores were able to recover germination ability after a period of seawater incubation once salinity concentrations decreased. Variable salinity conditions are a typical feature of sea cliffs where halophytes can grow. A salty culture medium also affects the development of the prothalli. Higher salinity of the substrate produced a significant decrease in gametophyte sizes, and affected the development of gametangia as well. Only males were found in a 0.98%-salt medium, and no gametangia were observed in cultures at 1.96%-salt medium. Recovered spores after one and a half month in high salinity concentration, 3.26%, were able, when transplanted to 0%-salt medium, to develop normal sexual prothalli, and this occurred in a shorter time than spores without any prior treatment. A. marinum can be considered as a halophyte fern that is able to germinate and grow in salty conditions. But it takes advantage of rainy periods that cause decrease of salt concentration, which seems to be important in particular for gametangia development and fertilization.     

药用植物乌蕨配子体发育的观察

药用植物乌蕨(Stenoloma chusanum(L.)Ching)配子体发育周期短而迅速,是一种研究蕨类配子体世代的生理生态和分子生物学等特征的理想试验材料.本文采用原生境腐殖土培养方法对乌蕨的孢子进行人工培养,观察并记录其孢子萌发和配子体发育的过程.结果表明:乌蕨孢子呈黄色,单裂缝,二面体形;孢子萌发为书带蕨型(Vittaria-type).丝状体4～6个细胞,有时具二叉分枝.片状体为长楔形,达6～8个细胞宽.成熟的原叶体为对称心形,裸露,发育为槲蕨型(Drynaria-type),并且具备一定的营养繁殖能力.原叶体上的假根为单细胞且不含叶绿体.精子器为盖裂开放,颈卵器短而直立.利用孢子培养的方法对乌蕨种苗进行人工繁殖,将为保护性开发利用这一重要药用资源奠定基础.     

阔叶鳞盖蕨配子体发育及卵发生的研究

采用显微镜和透射电镜对阔叶鳞盖蕨(Microlepia platyphylla)的配子体发育和卵发生过程进行了观察,以阐明其卵发生的细胞学机制,探讨其演化地位。阔叶鳞盖蕨孢子褐色,四面体形,具三裂缝,接种5~10d后孢子萌发,经丝状体和片状体阶段发育为心形原叶体,原叶体发育是铁线蕨型,通常为雌雄异株,精子器产生于不规则配子体的表面,颈卵器产生于心形原叶体生长点的下方,性器官是薄囊蕨型。卵发生研究表明,阔叶鳞盖蕨颈卵器产生于生长点下方表面细胞,经两次分裂形成了顶细胞、初生细胞和基细胞。其中初生细胞再经两次不等分裂产生卵细胞、腹沟细胞和颈沟细胞,此3个细胞通过胞间连丝紧密相连,随发育,腹沟细胞与卵细胞间形成了分离腔,但在孔区处始终通过胞间连丝相连,成熟卵细胞上形成了卵膜和受精孔,卵核表面产生了核外突,通过比较表明阔叶鳞盖蕨卵发生与蕨（Pteridium aquilinum）卵发生相似。     

Costs of sporophyte production in the moss,Dicranum polysetum

We investigated the cost of sporophyte production in the moss Dicranum polysetum both by examining patterns of growth and reproduction in unmanipulated shoots and by experimentally manipulating sexual reproduction.The estimated proportion of total carbon investment allocated to sexual reproduction in sporophyte-producing shoots over the study period was 74.8%. Unmanipulated shoots that aborted all sporophytes had a significantly higher growth in the top shoots than shoots that produced sporophytes. In sporophyte-producing shoots, total apical growth decreased proportionately with the number of sporophytes.Experimental prevention of sporophyte development resulted in significantly higher total apical growth of the gametophytes. Shoots where current perichaetia were lacking when marked had a mass increase in the top shoots similar to manipulated sporophytic shoots whereas sporophytic control shoots grew significantly less than these two categories.The difference between control shoots and manipulated shoots in the mass of vegetative apical growth was mainly because of different length increments whereas mass per unit length was similar between groups. The probability to reproduce sexually in the year after the manipulation, and the biomass allocated to this reproduction, were not affected by the experimental treatment.