High Frequency Sporophytes Regeneration from the Spore Culture of the Endangered Aquatic Fern <i>Isoetes coreana</i>

Using a mixed culture of megaspores and microspores from I. coreana, we established high frequency sporophyte regeneration system. After 20 days of culturing in MS basal medium, microscopic examination showed significant morphological changes and the microspore released numerous small vesicles into the culture medium. Megaspores also showed dramatic morphological changes during its incubation time in culture. The spore wall was cracked by the expansion of the megaspore (about 2 times increase in diameter). Simultaneously, brown spots were observed on the surface of the megaspores. The frequency of female gametophytes developing from immature megaspores cultured in MS basal liquid medium (pH 7) supplemented with 1 mgl-1 GA3 was 46%. However, these female gametophytes derived from megaspore only culture could not differentiate into sporophytes. The mixed culture of microspores and megaspores resulted in successful sporophyte regeneration. The highest frequency (12.3%) of green sporophyte regeneration from mixed spore culture occurred when the cultures were maintained at 25℃ under cool-white fluorescent light (40 μmol·m-2·s-1) with a 16 h photoperiod. Regenerated sporophytes were transferred to a test tube containing vermiculite and a sand mixture and left there until they had three leaves. After root growth and the fifth leaf had emerged, more than 95% of the regenerated sporophytes were successfully transferred to the soil and grown to mature plants. The sporophyte regeneration system established in this study could be successfully used for the restoration of the endangered aquatic species, I. coreana.

Laminaria gametophyte clone culture and its application in sporeling cultivation

In China, the total annual production of cultured Laminaria japonica reached half million tons in dry weight there few years. The routine sporeling culture technique conducted in the greenhouse took at least three and half months. In such a case, sometimes the sporelings died within a few days due to destructive diseases. In Order to overcome the mentioned problems, a new sporeling culture technique, the clone technique, is developed. The method includes three stead: (1) Gametophyte clone culture. The spores and the gametophytes are cloned in flasks under favorable environments. (2) Sporeling cul ture. Male and female clones are crushed and spread onto a frame to allow the gametophytes to attach to the substrata. The frames are cultured in tanks, and the sporophytes reach 1 cm in length within one and a half months. (3) Outgrowing of the plant. The frames are put in the open sea when seawater temperature decreased to 20℃. After one month, the sporelings are large enough to be transplanted. It is concluded that the clone technique has the following advantages: (1)Large amount of clones can be produced in a short period of time. (2) Clone seeding method makes it free from the biological rhythm, one can seed the plant anytime all the year round. (3) It takes only one and a half months to complete the process of sporeling cultivation in the greenhouse. At present, this technique is used in the breeding of new strains of Laminaria.

Karyological observations of Ulva linza chromosomes

Abstract Ulva linza is one of the species that causes green tides in the Yellow Sea,China.Due to the diffi culties in chromosomal preparation,the large numbers of chromosomes,and their relatively small sizes,there have been no reported studies on Ulva macroalgae chromosomes.The karyotypes and chromosomes in U.linza were observed after a series of treatments.The chromosomes were pretreated with 0.1%colchicine for 12 h and then mixed with enzymes.The samples were dropped from 30 cm height onto glass slides,which spread out the surface coat.These pretreatments were the optimal chromosomes preparation treatments.The prepared chromosomes were stained with 4',6-diamidino-2-phenylindole(DAPI),which is a fl uorescent probe sensitive and specifi c to DNA.The chromosome number in the haploid male and female gametophytes was n=9,and was 2 n=18 in the diploid sporophytes.The female gametophyte chromosomes were between 0.804 and 2.292μm in size,the male gametophyte chromosomes were between 0.917 and 2.916μm,and the sporophyte chromosomes were between 0.912 and 2.167μm.The relative sizes of the chromosomes were used to analyze the karyotypes of the female and male gametophyte chromosomes.The results provide a solid foundation for the basic technique that can be used to localize molecular markers of Ulva chromosomes.

The Effects of Sugars and Ethylene on Apospory and Regeneration in <i>Ceratopteris richardii</i>

In land plants, two distinct generations, gametophyte and sporophyte, alternate to complete the life cycle. Sporophytes undergo meiosis to produce spores, from which gametophytes develop. Gametophytes produce gametes, which participate in fertilization to produce the zygote, the first cell of the sporophyte generation. In addition to this sexual reproduction pathway, some fern species can undergo apospory or apogamy, processes that bypass meiosis or fertilization, respectively, to alternate between the two generations without changing the chromosome number. Apospory is inducible in the laboratory in various fern species simply by altering the sugar level in the media. In sporophytes induced to undergo apospory, sporophyte regeneration is also observed. The ratio of aposporous gametophytes to regenerated sporophytes varies, in a manner consistent with being dependent on sugar level. Whereas the sugar signaling pathway is yet to be elucidated in lower plants, in angiosperms it has been shown to play a regulatory role in controlling essential processes including flowering and embryo development, which give rise to the gametophyte and the next sporophyte generation, respectively. Here, we present evidence for the role of different sugar levels on the balance of apospory and regeneration in the fern Ceratopteris richardii. The demonstration of crosstalk between sugar signaling and the hormone ethylene signaling in angiosperms prompted us to test the effects of this hormone in combination with sugar on apospory vs. regeneration. These results provide insight into how a group of redifferentiating cells determines which generation to become and lay the groundwork for further analysis of this asexual pathway.

<i>Conocephalum conicum</i>(L.) Dumort: A Case of Unique Reproductive Biology

This paper reports spore-elater ratio per capsule in three populations of Conocephalum conicum collected from different regions of Jammu and Kashmir (Doda, Ladakh and Bhaderwah). Spore-elater ratio came out to be 0.40-0.43:1, far less than expected for Marchantialian taxa. The ratios thus obtained were compared with that present in herbarium specimen collected in 1958 from Kyushu. The ratios have remained constant since many decades, thereby indicating that the sex-ual reproduction has lesser role to play in the propagation of this species.

Phototropism in the Marine Red Macroalga Pyropia yezoensis

Phototropism is a response to the direction of light that guides growth orientation and determines the shape of plants to optimize photosynthetic activity. The phototropic response is present not only in terrestrial plants but also in water-living algae. However, knowledge about phototropism in Bangiophycean seaweeds is limited. Here, we examined the phototropic response of the red alga Pyropia yezoensis to elucidate the regulatory mechanism of phototropism in Bangiophyceae. When leafy gametophytes and filamentous sporophytes of P. yezoensis were cultured under directional light, phototropism was observed in the gametophytes. Conchosporangia on the sporophytes also exhibited phototropism. Phototropism was positive in the majority of gametophytes and conchosporangia but in some cases was negative. In addition, a strong phototropic response occurred under white light, whereas blue and red light elicited minor and no responses, respectively. This observation is in contrast with the phototropic response in terrestrial plants and several algae, in which blue light is responsible for positive phototropism. Surprisingly, the genome of P. yezoensis has no homologues of the photoreceptors for blue and red light, revealing differences in the regulation of phototropism between terrestrial plants and P. yezoensis . Studies on the phototropism in P. yezoensis could shed light on the evolutional divergence of phototropic responses in plants.

Inter-specific and Inter-generic Hybridization Compatibility of Eriobotrya Species (Loquat) and Related Genera

We studied the cross-compatibility among 91 inter-specific combinations and 21 inter-generic combinations in 7 Eriobotrya plants and 2 related genera(Raphiolepis indica Lindl. and Photinia serrulata Lindl.) using emasculation, bagging, and artificial pollination. Our results showed that28 of the 91 inter-specific combinations set no fruit, which means nearly 30% of the combinations were incompatible. In the remaining 63 combinations, most showed partial cross-compatibility, and a few showed complete cross compatibility. Eriobotrya plants were incompatible with plants from their related genera(R. indica Lindl. and P. serrulata Lindl.). Backcrossing produced 5 compatible combinations, which could set fruits and produce F1 progeny but only after embryo rescue. Fruit setting ratios varied among various species used as male or female parents.E. prinoides Rehd. & Wils., common loquat(E. japonica) and Eriobotrya × daduheensis, used as female parents resulted in an average fruitsetting ratio of 36.2%–58.2%. E. deflexa Nakai and its two forms, and E. elliptica Lindl. as female parents resulted in 2.9%–16.3% average fruitsetting ratio; however, the fruit set ratio was higher(22.4%–43.1%) if they were used as male parents. Failure of E. deflexa f. koshunensis Nakai × E.prinoides Rehd. & Wils. hybrids to set fruit could be attributed to sporophytic incompatibility.

Excitation energy distribution between two photosystems in Porphyra yezoensis and its significance in photosynthesis evolution

Comparative investigation on energy distribution between two photosystems were carried out in the sporo-phytes and gametophytes of Porphyra yezoensis. By performing 77 K fluorescence spectra, we suggested that there probably existed a pathway for energy transfer from PS II to PS I to redistribute the absorbed energy in gametophytes, while no such a way or at minor level in sporophytes. Electron transfer inhibitor DCMU blocked the energy transfer from PS II to PS I in gametophytes, but no obvious effects on sporophytes. These indicated that excitation energy distribution between two photosystems in gametophytes was more cooperative than that in sporophytes. These data in ontogenesis reflected the evolution process of photosynthetic organisms and supported the hypothesis of independent evolution of each photosystem.

Sporophytic control of anther development and male fertility by glucose-6-phosphate/phosphate translocator 1(OsGPT1) in rice

Coordination between the sporophytic tissue and the gametic pollen within anthers is tightly controlled to achieve the optimal pollen fitness. Glucose-6-phosphate/phosphate translocator(GPT) transports glucose-6-phosphate, a key precursor of starch and/or fatty acid biosynthesis, into plastids. Here, we report the functional characterization of Os GPT1 in the rice anther development and pollen fertility. Pollen grains from homozygous osgpt1 mutant plants fail to accumulate starch granules, resulting in pollen sterility. Genetic analyses reveal a sporophytic effect for this mutation. Os GPT1 is highly expressed in the tapetal layer of rice anther. Degeneration of the tapetum, an important process to provide cellular contents to support pollen development, is impeded in osgpt1 plants. In addition, defective intine and exine are observed in the pollen from osgpt1 plants. Expression levels of multiple genes that are important to tapetum degeneration or pollen wall formation are significantly decreased in osgpt1 anthers. Previously, we reported that At GPT1 plays a gametic function in the accumulation of lipid bodies in Arabidopsis pollen. This report highlights a sporophytic role of Os GPT1 in the tapetum degeneration and pollen development. The divergent functions of Os GPT1 and At GPT1 in pollen development might be a result of their independent evolution after monocots and dicots diverged.