Clematis species are perennial woody or herbaceous vines that are popular medicinal and ornamental horticul-tural plants. The seeds of six species were collected from five sites, and their germination characteristics and dormancy types were investigated. Seeds of all species had a typical achene morphology of a seed pod with a tail. Small, rudimentary embryos (0.14 to 0.29 E:S ratio) were observed, and the seeds showed sufficiently good water absorption (53.9-72.4%). Germination occurred within 30 days for C. apiifolia (light and dark conditions, 66.3% and 90.5%, respectively), C. heracleifolia var. urticifolia (77.0 and 92.5%), C. heracleifolia (37.3 and 88.3%), and C. serratifolia (1.0 and 96.5%). Germination was promoted using gibberellin treatments. Furthermore, the seeds of C. terniflora and C. trichotoma broke dormancy with short-term cold temperature stratification, and the em-bryos developed and germinated at warm temperatures. We concluded that the seeds of C. apiifolia, C. her-acleifolia var. urticifolia, C. heracleifolia, and C. serratifolia are morphologically dormant whereas those of C. terniflora and C. trichotoma are morphophysiologically dormant. This study provides seed propagation and dormancy information for six Clematis species native to Korea.

Athyrium acutipinnulum Kodama ex Nakai is an edible fern that is native to Ulleung island and Gangwon Province (Taebaek Mountain), South Korea. A. acutipinnulum has high nutritional value and is rich in protein, carbohydrates, vitamins, minerals, and nicotinic acid. However, the cultivation of A. acutipinnulum is generally carried out on a small scale. In addition, the current rhizome propagation method is labor intensive and inefficient with regard to root growth-point establishment. The present study was conducted to improve the efficiency of the A. acutipinnulum cultivation method via in vitro culture and synthetic seed technology. To select an effective medium for in vitro proliferation, the concentrations of mineral salts, sucrose, activated charcoal, and total nitrogen in Murashige and Skoog (MS) media were adjusted. The 1 MS medium (3% sucrose, 60 mM total nitrogen, 0.8% agar, pH 5.8), had the highest gametophyte proliferation. Sporophyte formation and growth were outstanding in the mixed substrate including horticultural substrate and decomposed granite 2:1. The production of SFG (synthetic seeds using fern gametophyte) using the proliferated gametophyte led to a percent of sporophyte formation of 76%. Plant growth regulators were added in SFG in order to increase the percent of sporophyte formation. Gibberellin acid (GA 3 ) added SFG showed an increase in the percent sporophyte formation to more than 90%. The present study explored the optimal growth medium conditions for A. acutipinnulum gametophytes to use for SFG. SFG were successfully produced and their conditions for the massive sporophyte formation were confirmed.

Cryopreservation is an important technique used in the conservation of various plant tissues. This study proposes a cryopreservation method for the long-term conservation of eastern bracken fern gametophytes ( Pteridium aquilinum var. latiusculum ). Encapsulation–dehydration of the gametophytes was performed, and the exogenous sucrose and abscisic acid (ABA) preculture conditions were investigated. Gametophytes are sensitive to dehydration and drying, and the following treatment conditions were applied: encapsulation by alginate containing 0.75 M sucrose, 18-h loading treatment with 0.75 M sucrose, and 6-h drying treatment. The survival rate following cryopreservation was determined. The water content of < 27.5% in the alginate beads after dehydration and drying was found to be appropriate for ensuring survival. Additionally, performing an exogenous sucrose and ABA preculture was essential before encapsulation to achieve a survival of ≥ 90%. The high stress induced by cryopreservation and exogenous preculture regulated the expression of PaSuSy , PaLEA14 , and PaABI1b and the endogenous ABA content. In eastern bracken gametophytes, ABI1 appears to be a negative regulator of ABA signaling. These results indicate that the encapsulation–dehydration method is effective for the long-term conservation of eastern bracken fern gametophytes, and exogenous preculture alleviates abiotic stress and increases the survival rate.

Matteuccia struthiopteris (L.) Tod. is one of the edible ferns consumed globally. However, efficient M. struthiopteris propagation methods have not been developed, and current cultivation methods based on rhizomes are labor-intensive. The aim of the present study was to investigate the optimal in vitro and ex vitro conditions for the mass propagation of M. struthiopteris through SFG (synthetic seeds using fern gametophyte) technology. In vitro gametophyte proliferation experiments were carried out using 0.2 g of homogenized gametophyte cultured in modified MS medium (full-strength MS, 3

Background Liquid suspension culture efficiently proliferates plant cells and can be applied to ferns because it rapidly increases the fresh weight of gametophytes. This study assessed gametophyte proliferation and sporophyte production of Pteridium aquilinum var. latiusculum using a suspension culture method. Results The growth curve linear phase of gametophyte cells was confirmed between 9 and 18 days of culture, and the subculture cycle was determined to be 2 weeks. A double-strength MS medium (fresh weight, 18.0 g) containing 2% sucrose and NH 4 + :NO 3 − (120 mM, 40:80) was found to be the optimal liquid medium. Gametophytes obtained after suspension culture for 18 days did not normally form sporophytes in an ex vitro soil environment. However, this issue was resolved after changing the culture type or extending the culture period to 6 weeks. A short suspension culture period increased the fresh weight of fragmented and homogenized gametophytes but yielded numerous relatively immature gametophytes (globular forms of branching gametophytes, BG). Furthermore, differences in gametophyte morphogenesis and development were indicated by changes in endogenous phytohormone content. BG with immature development exhibited high accumulation of zeatin, jasmonic acid, and salicylic acid, and relatively low levels of abscisic acid and indole-3-acetic acid. The immature development of gametophytes directly affected sporophyte formation. Conclusions This study maximized the advantages of liquid suspension culture using eastern bracken gametophytes and provides data to resolve any associated issues, thus facilitating efficient bracken production.

Desiccation conditions and storage temperature are crucial factors for the successful long-term storage of fern spores. This study aimed to determine the optimal desiccation conditions and storage temperatures for the long-term survival of eastern bracken (Pteridium aquilinum var. latiusculum) spores. The effect of different storage temperatures (25°C, 4°C, −20°C, −80°C, and −196°C for up to 48 weeks) on spore germination and sporophyte development was investigated in collected spores. The germination percentage remained constant in all spores, except in those stored at −20°C and 25°C. Spores stored at −20°C had the lowest germination percentage and were almost completely nonviable. The collected spores were desiccated for one week at various relative humidity (RH) levels equilibrated using saturated salt solutions. The water content was measured in desiccated spores, and the spores were stored in liquid nitrogen for up to 48 weeks. The water content of the spores desiccated at 15–55% RH was 5.0–5.4%, and their germination percentage remained constant with storage time. Desiccation at 94% RH increased the spore water content to 9.5% and decreased the germination percentage. The relatively high water content in spores was not suitable for their long-term storage. Long-term survival of the spores can be achieved by desiccating the spores at low RH to adjust the spore water content to 5% prior to their storage at low temperature.

The formation and pollution of particulate matter (PM), a side effect of rapid industrialization and urbanization, is considered a global issue. However, various plant species are able to effectively capture and reduce atmospheric PM concentrations. We investigated the indoor growth and morphology of 21 indigenous Korean evergreen species at low light intensities to ascertain their ability to reduce PM of aerosol particles in a closed acrylic chamber. The decrease in PM mass concentration differed significantly across species, with a significant correlation (8 h; p < 0.001). The reduction in the mass concentration of PM differed with particle size and across species. The highest reduction of PM2.5 occurred after 8 h with Dryopteris lacera (86.8%), Ilex × wandoensis (84.9%), Machilus thunbergii (84.3%), and Rhododendron brachycarpum (84.0%). Reduction of PM10 after 8 h was highest with Cephalotaxus harringtonii (98.3%), I. × wandoensis (98.5%), M. thunbergii (98.5%), and R. brachycarpum (98.3%). Plant morphological characteristics (category, plant height, leaf shape, leaf area) and relative humidity were closely related to the decrease in PM mass concentration. In conclusion, our findings can be used to identify Korean plant species that can reduce PM concentration and are suitable for indoor use.

This study proposed the production of gametophyte-derived synthetic seeds and a practical plug seedling method for eastern bracken. Synthetic seeds were produced by mixing fragmented gametophyte and algi-nate matrix. The gametophyte regenerated regardless of the syringe tip size and the injection molding type employed, to successfully form the sporophyte. The synthetic seeds were successfully stored short-term under cold storage (4 degrees C). The storage period influenced the percentage of sporophyte formation, which was 70.8% or more until day 7, but decreased to 56.9% or less after day 14 of storage. However, the storage period of 7 days is sufficient for short-distance transport. The development of gametophyte-derived synthetic seeds improves the logistics of transport and handling ferns, and it enables establishment of plug seedling cultiva-tion. (c) 2021 SAAB. Published by Elsevier B.V. All rights reserved.

Selaginella tamariscina is a medicinal plant that contains a variety of plant secondary metabolites; however, it is currently being collected indiscriminately from its native habitats. Hence, we have developed an efficient propagation method for S. tamariscina. Explants grown in vitro were cultured in Murashige and Skoog medium of various strengths (1/16–2x), and the highest number of sporophytes (65.7) were obtained with 1/4x MS medium. Culturing explants at various lengths (3–12 mm) for 12 weeks indicated 12 mm as the most appropriate size for sporophyte propagation. We then evaluated various concentrations of individual components, sucrose (0–5%), total nitrogen (7.5–30 mM), nitrogen ratio (3:0–0:3), and agar (0.6–0.8%), in the 1/4x MS medium for explant growth for 12 weeks. The maximum number of sporophytes were formed in media containing 3% sucrose, 15 mM nitrogen, and 0.6% agar, with a nitrogen ratio of 1:2. The propagated S. tamariscina was then acclimatized in a controlled environment to improve survival in an external environment. These results demonstrate the effective conditions for in vitro mass propagation of S. tamariscina, finding that methods utilizing sporophytes were more efficient than conventional propagation methods and yielded numerous plants in a short period.

The effect of different conditions for in vitro proliferation, and soil conditions for sporophyte formation, were studied in Onoclea interrupta (Maxim.) Clang & P. C. Chiu. The gametophytes produced from in vitro-germinated spores were then cultured for eight weeks on Murashige and Skoog (MS) medium at quarter-, half-, full-, and two-fold strength, and full-strength Knop medium. Maximal fresh weight gains (10.2 g) and excellent rhizoid and gametophyte morphogenesis were obtained with full-strength MS medium. The effects of sucrose concentrations (0, 1, 2, 3, 4%) and activated charcoal (0, 0.2, 0.4, 0.8%) in full-strength MS medium, and the effect of different N concentrations (30, 60, and 120 mM) were also studied. Sucrose at 1% led to the highest gametophyte weight (4.2 g). Activated charcoal did not affect weight. Nitrogen with an NH 4 . to NO, ratio of 1 : 2 at 30 mM led to good weight gain (3.5 g). Sporophyte formation occurred on all researched soil substrates after 12 weeks. Maximum sporophyte production (405 sporophytes) occurred in mixed soil containing horticultural substrate and decomposed granite in a ratio of 2 : 1 (v/v). These sporophytes grew well in terms of leaf length, number of leaves, and number of roots.

Information on the optimal conditions to promote the germination of Lamprocapnos spectabilis (L.) Fukuhara seeds is limited; consequently, this study was conducted to establish the requirements to break their dormancy and promote germination. The selected seeds were morphophysiologically dormant and had not begun embryo development. To study the dormancy breaking and embryo development processes, seeds were subjected to constant or changing temperature treatments during moist stratification. High temperature and humidity conditions resulted in vigorous embryo growth, with the longest embryos occurring after a month of incubation at 20 ℃. At 4 ℃, the seeds required incubation period of at least three months to germinate. Embryo growth and germination were greater with changing high and low temperatures than under a constant temperature, and changing temperatures also considerably changed the endogenous hormone levels, embryo development, and germination. Bioactive gibberellin (GA) content was higher in the seeds incubated at 20 ℃ for one month, then at 4 ℃ for two months. The content of endogenous abscisic acid in the seeds subjected to the same treatment decreased by 97.6 %, compared with that of the untreated seeds. Embryo growth and seed germination require changing high and low temperatures; however, exogenous GA3 could substitute for high temperatures, as it also causes accelerated germination. In this study, the seeds of L. spectabilis were identified as an intermediate simple type, a sub-level of morphophysiologically dormant seeds.

Pteris fauriei Hieron. was first reported as a new fern species for South Korea in 2014. Since then, propagation information for this species has not been clearly documented. Thus the aim of our study was to determine the in vitro spore germination conditions of P. fauriei and to suggest an efficient propagation method using mechanically fragmented gametophytes. Spore germination was tested under different light spectra (cool white fluorescent light (CWFL), red and blue light), temperature (15, 20, 25, and 30 degrees C), and culture medium (Knop and one-eighth, quarter-, half-, full-, and double-strength Murashige and Skoog (MS)) conditions. Optimal conditions for the best germination (96.1%) were Knop medium, 25 degrees C, and the CWFL. Blue light, low temperature, and double-strength MS medium conditions inhibited spore germination. Sporophyte production by applying the mechanical fragmentation method was done using gametophytes obtained from spore germination. This method produced a maximum of 114.5 sporophytes in full-strength MS medium in vitro. Therefore, we suggest mass production methods for P. fauriei seedlings, which can be easily implemented in the in vitro systems.

Bracken fern (Pteridium aquilinum var. latiusculum (Desv.) Underw. ex A. Heller) has long been grown industrially in South Korea. Conventional propagation methods, including planting rhizomes and in vitro seedling culture, are labor intensive and expensive, and thus not commercially suitable. We aimed to develop a system to produce synthetic seeds using fern spores (SFS). Synthetic seeds were prepared by mixing bracken spores and alginate matrix. Spore germination and gametophyte and sporophyte growth and development from SFS proceeded normally. Spore density affected gametophyte and sporophyte numbers. SFS prepared using cold (4 °C) long-term storage spores (even 7-year-old spores) could effectively form sporophytes. The highest germination was observed at 25 °C. Soaking-treated SFS successfully formed sporophytes, even after 30 days of storage at 4 °C; indeed, sporophytes formed even after five days of storage at 25 °C during transport conditions. SFS were sown in plug trays for commercial use. Young sporophytes grown from plug seedlings were greenhouse cultivated, and transplanting within eight weeks was effective for root growth and growing-point formation. Developing synthetic seeds is a feasible solution for facilitating efficient transport and the handling of small-sized fern spores; furthermore, this SFS technology provides the basis for fern seedling culture and fern spore industrialization.