In vitro germination and propagation of a threatened medicinal orchid,Cymbidium aloifolium(L.) Sw.through artificial seed

Objective:To study the in vitro germination and plantlet regeneration from artificial seeds of Cymbidium aloifolium(C.aloifolium),a highly threatened medicinal orchid of Nepal.Methods:Artificial seeds were produced in vitro by encapsulation of protocorms with 4%sodium alginate and 0.2 mol/L calcium chloride solution.In vitro germination and plantlet regeneration of the artificial seeds were tested by culturing them on different strength of Murashige and Skoog(MS) liquid media(0.25,0.5 and 1.0) and MS liquid medium supplemented with 0.5 mg/L benzyl amino purine and 0.5 mg/L naphthalene acetic acid.Freshly produced artificial seeds were stored up to 28 d at 4 ℃.In order to check the viability,storage artificial seeds were treated with five different sterilization techniques(T_1 T_2,T_3 T_4,T_5) and inoculated on full strength(1.0) of MS liquid medium after each 7 d of interval upto28 th days.Results:The highest percentage of germination(100%) of artificial seed was obtained on quarter(0.25),half(0.5) and full(1.0) strength of MS liquid medium.Experimentally,full strength of MS liquid medium was more effective for earlier seedling development of C.aloifolium.Artificial seeds were successfully stored at 4 ℃ till 28 th days.Treatments T_1 and T_2showed 97.5%viability of storage artificial seeds and hence considered as the most effective sterilization techniques to recover the plant from storage artificial seeds.Plantlets developed from artificial seeds were successfully acclimatized in potting mixture containing cocopeat,litter and sphagnum moss with 85%survival rate.Conclusions:The present study revealed that artificial seeds are the good alternative explants for in vitro mass propagation and short term conservation of C.aloifolium.

Efficient Plant Regeneration of Cymbidium aloifolium (L.) Sw., a Threatened Orchid of Nepal through Artificial Seed Technology

Artificial seed technology is a method of considerable potential for mass propagation and conservation of rare, endangered and threatened species. In the present investigation, artificial seeds were obtained through encapsulation of three weeks old protocorm (3.0 ± 1.0 mm diameter) of Cymbidium aloifolium with calcium alginate. Artificial seeds were cultured on liquid Murashige and Skoog (MS) or Knudson (Kn C) medium at different strength (×1.0, 0.5, 0.25) and full strength (1.0) of both media supplemented with 0.5 mg/l 6-benzyl aminopurine (BAP) and 0.5 mg/l α-naphthalene acetic acid (NAA). Full strength of MS medium without plant growth regulators was found to be the most favourable condition for efficient plantlet regeneration of C. aloifolium (9.83 shoot and 2.66 roots per culture). The storage potential of artificial seed was tested at 4°C and room temperature (RT, 21°C ± 2°C) for up to 90 days on both media and found 83.33% viability at 4°C storage on MS media. Eighty five percent of plantlets regenerated from artificial seeds culture were successfully hardened in a potting mixture of cocopeat, clay and sphagnum moss (2:1:1). Hence, the methodology can be used for propagation and conservation of C. aloifolium through artificial seed system.

Study on Artificial Seeds of Plants

The establishment and development of artificial seed technology is to quickly reproduce excellent varieties or hybrids,which can be applied to hybrid generation seeds produced by the three-two line method.For some varieties that are difficult to propagate with seeds or plant species with unstable genetic traits and poor fertility,artificial seed technology can also be used for mass reproduction.In particular,some new plants created through genetic engineering,such as somatic hybrids or transgenic plants,can be propagated or maintained by artificial seed technology.In addition,artificial seed technology can be used for the maintenance and rapid propagation of virus-free seedlings.Compared with ordinary test tube seedlings,artificial seeds have low cost,convenient transportation,and to a certain extent reduce vitrified seedlings.In particular,the production of artificial seeds does not occupy a large amount of soil.It can be produced all year round.Therefore,the research on artificial seeds has developed rapidly in the world.

Regeneration of Cytologically Stable Plants Through Dedifferentiation, Redifferentiation, and Artificial Seeds in Spathoglottis plicata Blume.(Orchidaceae)

Spathoglottis plicata Blume. is a horticulturally important vulnerable ground orchid with beautiful flowers blooming round the year. Highfrequency protocorm-like body(PLB) formation was established via callus culture from vegetative tissues of in vitro germinated seedlings of S.plicata. Media containing MS salts and Gamborg's B5 vitamins supplemented with 1.0 mg·L^(-1) 2,4-dichlorophenoxyacetic acid(2,4-D), 3.0 mg·L^(-1) α-naphthaleneacetic acid(NAA), 1.0 mg·L^(-1) kinetin(KIN), and 10%(v/v) ‘Aloe vera gel'(Av G) were effective in fragile calli induction. A maximum of(22.3 ± 0.52) PLBs were induced from about 250 mg callus within 45–55 days in the presence of 2.0 mg·L^(-1) NAA and 3.0 mg·L^(-1) 6-benzylaminopurine(BAP). Briefly, 3.0% sodium alginate was found to be most suitable for the formation of an appropriate shape and good germination rates(86.7%)of artificial seeds. Out of three different temperatures(4, 15, and 24 °C), the best result was achieved at 4 °C with 66.7% germinability even after90 days of storage. Plantlets were acclimatized with 86.6% survival rate and 76.3% of these plants produced flowers within 12–15 months of field transfer. Chromosomal studies revealed cytological stability of all regenerants containing 2 n = 40 chromosomes as in the parental plants.The present protocol can be applied reliably for the purposes of large-scale commercial propagation and short-term conservation of this orchid.

Artificial Plant Seeds and Their Application

The artificial plant seed is a new technology developed on the basis of the in vitro plant culture technology.As a high-tech achievement with great development potential and economic value in the 21^(st) century,artificial plant seeds have great potential advantages.This paper briefly introduced artificial plant seeds,their composition and advantages,the preparation of artificial plant seeds and the application of artificial plant seeds.

Encapsulation of Immature Somatic Embryos of Coffea arabica L.for in Vitro Preservation

The present study aimed to develop a protocol for somatic embryogenesis and encapsulation of coffee embryos(Coffea arabica L.),for the conservation of genotypes with characteristics of commercial interest.Somatic embryos were induced from leaf explants in Murashige and Skoog medium(MS)supplemented with 1 mg·L^(−1) of 2,4-dichlorophenoxiacetic acid(2,4-D)combined with 2 mg·L^(−1) of benzyladenine(BA).Somatic embryos(SE)at the globular stage were encapsulated in a sodium alginate matrix;two treatments were tested:MS+5 mg·L^(−1) BA+1 mg·L^(−1) NAA+3%(w/v)alginate,and MS+7 mg·L^(−1) BA+5.7 mg·L^(−1) indoleacetic acid(IAA)+3%(w/v)alginate.Alginate was complexed with 100 mM calcium chloride(CaCl_(2)).Viability of the encapsulated SE was determined by staining with 0.01%fluorescein diacetate(FDA)after 0,15,30,and 45 days of storage at 4℃.Embryo viability was 100%in both treatments.

Regulation Culture on Cytological, Biochemical and Physiological Characteristics of Somatic Carrot Embryos

Cytological,biochemical and physiological charcteristics of quiescent,high-vigor carrot somatic embryos obtained with the use of regulation culture were studied. As shown in cytological observation, quiescent somatic embryos obtained by regulation culturing for 70 days displayed small compacted cells with thick wall, dense cytoplasm and large amount of starch granules. Biochemical tests revealed a starch content and a soluble sugar content 1.9 and 1.7 times respectively as much as those of the control embryos(normal culture) while physiological study demonstrated that the respiration was maintained stally at a very low level,only 1/4 that of the control at the end of the 70-day period. These results provide further evidences that regution culture so after the morphology, biochemistcy, physiologr and metabolism of somatic embryos that they became quiescent,highly synchronized, and had their roots ungrown, a sate facilitating safe storage.

Model Analysis of Radar Echo Split Observed in an Artificial Cloud Seeding Experiment

An artificial cloud seeding experiment was performed over the Japan Sea in winter to show how massive seeding could be effective to mitigate heavy snowfall damage.The results showed that 20 min after cloud seeding,a portion of the radar echo beneath the seeding track was weakened to divide the radar echo into two parts.In order to analyze the results,a numerical simulation was conducted by using the Weather Research and Forecasting model verion 3.5.1.In this simulation,the seeding effects were represented as phenomena capable of changing rain particles by accreting cloud ice and snow to form graupel particles and by changing cloud liquid water to snow particles.The graupel particles fell rapidly,thus temporarily intensifying the rainfall,which subsequently decreased.Therefore,the weakened radar echo in the field experiment is deemed to have been caused by the increase in rapidly falling graupel particles.

An Example of Canal Formation in a Thick Cloud Induced by Massive Seeding Using Liquid Carbon Dioxide

The purpose of this experiment is to show that massive cloud seeding is effective in mitigating the damage caused by heavy snowfall. In order to show its effect, we attempted to form a canal in a thick convective cloud by massive seeding, and left the parts that were not influenced by the seeding as a reference to show that the canal was formed by the massive seeding only. The seeding was carried out by using an aircraft. The seeding rate and air speed of the aircraft were 35 g s-1 and 115 m s 1, respectively. The flight course for seeding was selected to be parallel to the wind direction to ensure that the dispersed liquid carbon dioxide did not influence both sides of the course. The results show that a part of the radar echo observed from onboard beneath the seeding track was weakened and divided the radar echo into two parts 20 minutes after the cloud top and the bottom were seeded, and distribution of rainfall rate on the ground from the target cloud was confirmed to be divided into two parts 24 minutes after the seeding. The target cloud was torn along the seeding track, and we could see the sea surface through the break in the cloud. Canal formation occurred in the cloud along the seeding track. Clouds and snowfall were left on both sides of the canal. The results show that supercooled liquid cloud particles along the seeding track evaporated to form larger precipitable particles which grew and fell rapidly.