• Horticultural Science & Technology
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• v.29 no.5
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• pp.494-499
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• 2011

Cold pretreatment, washing medium and composition of nutrient media may have marked effects on microspore embryogenesis. When microspores isolated from radish (Raphanus sativus L. cv. Gwanhun) flower buds were washed with Nitsch & Nitsch (NLN) medium liquid medium containing $130g{\cdot}L^{-1}$ sucrose (NLN-13), yields of microspore-derived embryos were greater than when using B5 liquid medium containing $130g{\cdot}L^{-1}$ sucrose. Microspore viability is known to decrease rapidly with storage; however, in this experiment, microspore viability was maintained for 24 h at $4^{\circ}C$ without media. Among the various medium concentrations used ($0.25{\times}$, $0.5{\times}$, $1.0{\times}$, $2.0{\times}$, and $4.0{\times}$ NLN liquid medium), $0.5{\times}$ NLN liquid medium induced the most efficient formation of microspore-derived embryos. In addition, microspore-derived embryos yields were greater when microspores were cultured in $0.5{\times}$ NLN liquid medium supplemented with $0.25{\times}$, $0.5{\times}$, and $1.0{\times}$ NLN microelements, compared to medium not supplemented with microelements. In this study, the highest yield of microspore-derived embryos was observed when the microspores derived from flower buds were washed using NLN-13 liquid medium and then cultured on $0.5{\times}$ NLN liquid medium supplemented with $0.25{\times}$ NLN microelements, followed by incubation at $25^{\circ}C$ for 30 days.

• Journal of Plant Biotechnology
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• v.38 no.1
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• pp.30-41
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• 2011

The influences of the agitation as well as the addition of medium during culture on the production of embryos were invested in isolated microspore culture of hot pepper (Capsicum annuum L.). When the culture medium was added during initial liquid culture step of liquid-double layer culture, the embryo yield and quality greatly increased. The most effective time point for medium addition was 5 days after the culture commenced. On the other hand, the effect of medium addition at later double layer culture step in liquid-double layer culture on the embryo production was less compared to that of medium addition during the initial liquid culture step. Agitating the culture for 1 week during later double layer culture step in liquid-double layer culture effectively increased the production of normal cotyledonary embryos. In the case of liquid culture, agitating the culture for 1 week from 7 days after the culture commenced was also effective for embryo development. However, when the total agitation time was longer (2 to 3 weeks) during liquid-double layer culture or liquid culture, the embryos developed abnormally in both cases. The normal cotyledonary embryos obtained in this study successfully developed to plants when transferred to regeneration media. These regenerated plants were either diploid or haploid, and there was a difference in the number of chloroplasts between guard cells of diploid and haploid. These results can be used as an important data for developing an efficient microspore culture system with high quality embryo production in hot pepper.

• Korean Journal of Plant Resources
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• v.18 no.2
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• pp.302-308
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• 2005

This study was carried out to obtain the basic informations on the characteristics of gametophytes formation and embryo developement in Dicentra spectabilis. Microspore mother cells developed from archesporial cells, start meiosis when flower bud length reaches around 1 mm, formed tetrahedral type tetrad. The 4 microspores were separated. They were developed to male gametophytes, respectively. Megaspore mother cells were observed when flower bud length was $4{\sim}5\;mm$. The developemental type of megaspore was polygonum and embryo sac was amphitropous. Three large and distinctive antipodals did not degenerated and remained after embryo sac was developed. When the male and female gametophytes was fully developed, the length of stamen and style was very similar or stamen was shorter about 0.5 mm than that of style. This result indicates that self-fertilization can be occurred in this species. After fertilization, developing zygotic embryos showed various stages of development from globular to cotyledonary embryos, and zygotic embryo in seed scattering time seemed to have an early cotyledonary stage.

• Proceedings of the Korean Society of Plant Biotechnology Conference
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• 2002.04b
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• pp.183-183
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• 2002

• KOREAN JOURNAL OF CROP SCIENCE
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• v.53 no.2
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• pp.150-155
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• 2008

This experiment was carried out to investigate the effect of proton ion and gamma-ray irradiation on microspore culture of the flower buds of $M_2$ generation in winter type of Brassica napus L. ssp. oleifera. The seeds of three rape varieties, 'Halla', 'Naehan' and 'Tammi' were pretreated with proton ion and gamma-ray 400 Gy and 600 Gy, respectively. When microspore culture techniques were used, embryogenesis was increased in some varieties by proton ion and gamma-ray irradiation treated flower buds of $M_2$ generation than control. In genotypes 'Naehan' showed the highest embryo production frequency, but 'Tammi' showed lowest embryo production frequency. Some of the embryoids developed directly into plantlets, whereas others developed abnormally multilobe. Plants were regenerated and successfully acclimatized in pots.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.46 no.6
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• pp.443-448
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• 2001

This study was carried out to obtain the basic informations on the characteristics of gametophytes formation and microspore germination in Astragalus membranaceus Bunge. Pollen mother cells passed through meiosis when the flower bud length reaches around 3.5 mm, thus creating the tetrad when it is 4.0 mm long. Pollen attains full growth when the bud is about 10.0 mm long and the anther is found to dehisce when the length of tate bud reach around 12.0 mm. Embryo sac develops at a similar speed as pollen did and it attains its full growth when the bud is about 10-12 mm long. After being stored at 4$^{\circ}C$ or -4$^{\circ}C$, the pollen maintained its germination ability to almost full extent by the 30th day after store. However, the germination rate at room temperature (23~28$^{\circ}C$) decreased below 3％ by the 3rd day of storage and so did the germination speed.

• Korean Journal of Plant Tissue Culture
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• v.26 no.3
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• pp.149-156
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• 1999

In order to investigate the effect of low temperature pretreatment on pollen dimorphism and embryo formation in anther culture of Platycodon grandiflorum, the anthers with microspore at the uninucleate stage were cultured on Murashige and Skoog medium supplemented with 0.5mg/L NAA and 1.0 mg/L BA. The low temperature pretreatment have clear effect on the frequencies of S pollen grains, symmetrical binucleate microspores (B type of S pollen), multinucleate and multicelled pollen grains. Especially, after low temperature pretreatment at 8$^{\circ}C$ for 5 days increased the frequency of S pollen grain (20.6%) in vivo. In addition, the highest frequency of callus induction (54.9%) and embryo formation (9.9%) were obtained from the anther pretreatment at 8$^{\circ}C$ or 5 days. Three distinct pathways could be recognized in the androgenesis, one involving mainly the vegetative cell, the second starting with the vegetative and the generative cell, respectively, and the third accompaning with two equal vegetative type cells in the pollen grains.

• Korean Journal of Plant Tissue Culture
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• v.27 no.5
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• pp.411-413
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• 2000

The ability of microspores or young pollen grains (male gametophytes) to undergo developmetal switch to embryogenic (sporophytic) pathway exemplifies the concept of totipotency as applied to haploid posmeiotic cells. As a first step pollen is devoid of positional information provided in situ by the intact anther - by isolation and cultivation in vitro in artificial media. This is inevitably accompanied by some degree of stress response in microspore/pollen. It has been shown in both monocots and dicots that intentional stress treatment (mostly starvation or heat shock) greatly stimulates embryo induction rate. Using transgenic sHSP antisense Nicotiana tabacum we show that expression of small heat shock proteins is an integral part of successful embryo and later haploid plant production from pollen grains. Our recently published data show that sHSP chaperone function is optimal in the absence of ATP.

• Journal of Plant Biotechnology
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• v.32 no.1
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• pp.37-44
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• 2005

Microspores were isolated from pepper (Capsicum annuum L.) anthers by using a micro-blender and cultured in modified NLN medium at $25^{\circ}C$. The influence of pretreatment period at $32^{\circ}C$, adding the 2-hydroxynicotinic acid to a pretreatment medium, and co-pretreatment anthers with microscopes on the induction of embryo were examined. Globular and torpedo embryos were observed from 3 weeks after culture. Embryo development was not synchronized within culture. After 4 weeks in culture, in addition to globular and torpedo embryos, cotyledonary embryos were observed. Normal cotylodonary embryos developed into plantlets when transferred to a solid hormone free B5 medium containing $2\%$ sucrose. Embryo yields were significantly higher after 1- and 2-day pretreatment at $32^{\circ}C$. However the development of embryo ceased at the globular or heart stage. In contrast, embryo yields were lower after 3- to 6-day pretreatment at $32^{\circ}C$ and embryo developed at the cotyledonary stage. After adding the 2-hydroxynicotinic acid to anther pretreatment solution, embryo yields were slightly increased. However most embryos occurred were at the globular or heart stage. Co-pretreatment of microspores with anthers was deleterious for embryo induction and development. AS far as we know, this is the first report of success in obtaining high frequency of embryogenesis and plantlets formation from isolated microspores of pepper. Although the culture conditions have to be optimized further, this promising microspore culture system can be used for genetic transformation, selection for dominant and recessive traits as well as for the production of homozygous doubled haploid plants.

• Journal of Korean Society of Forest Science
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• v.3 no.1
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• pp.1-4
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• 1963

Present experiment has been carried out in order to make clear the abnormalities of the female gametophyte formation and its relation to fertility, using the short-style of F. koreana, the results of which are summarized as follows : (1) Anatropous ovule has single integument with thick cell-layer and tiny nucellus consisting of nucella-epidermis and megaspore mother cell. (2) Meiotic division of megaspore mother cell takes place around middle or latter part of March, while that of microspore mother cell occurs from the end of September to the beginning of October. (3) Megaspore mother cell stage is long, and ranges from October to March next year. (4) Formation of mature embryo sac is not completed until the beginning of May, approximately one month after blooming. (5) Normal embryo sac is rare, most of the nucellus being devoid of embryo sac.