외래단백질의 생산을 위하여 백합화분을 이용한 일시발현체계를 개발하였다. 충분히 성숙단계에 도달한 백합의 수술로부터 백합화분을 수집한 후 이들은 40-100 μm 크기의 aluminum oxide 입자와 함께 강하게 교반시켰다. 결과적으로 표면에 상해를 지닌 화분립에 대하여 erythropoietin(EPO) 유전자 DNA를 포함한 식물발현벡터(pBI-EPO)를 지니는 Agrobacterium tumefaciens LBA4404 세포들을 섞어 진공침윤을 수행하였다. Southern blot 및 cDNA blot hybridization의 분석 결과 EPO DNA의 화분 genomic DNA 삽입과 EPO mRNA의 생성이 각각 성공적으로 확인되었으며 이러한 결과는 Agrobacterium을 이용한 형질전환화분이 신속하고 편리한 단백질공장으로서 이용될 수 있음을 시사하고 있다.

캘러스, 반수체, picloram, 전처리양식을 규명할 목적으로 배양효율에 미치는 화분발육 단계와 온도전처리가 미치는 영향에 대한 실험결과는 다음과 같다. ‘Dreamland’의 감수분열중인 약을 현 미경적방법으로 검경하여 본 결과 화뢰의 길이가 23.0-24.9mm는 tetrad 단계이었고 25.0-26.9mm는 uninucleate 단계, 27.0-28.9mm에서는 binucleate 단계 임을 확인할 수 있었으며, 저온 및 고온 온도처리 결 과는 전체적으로 캘러스 및 배 발생율은 저조하였고 특 이한 점은 발견할 수 없었다. Microgametogenesis 단 계별로 약배양 효율을 조사한 결과, late uninucleateearly binucleate 단계인 23.0-28.0mm에서 17.8% callus가 유도되었고, 식물체 재분화율도 6.7%로 나타 나 적정 배양 stage임을 확인할 수 있었다. 유기된 캘 루스를 picloram과 zeatin을 혼용한 MS 배지에 이식 하여 25oC에서 배양하여 식물체를 얻었다.

The complex analysis of male reproductive system of vines and fruit-tyees growing in zone of the Armenian Nuclear Power Plant (ANPP) is realized. The obtained data demonstrate that the homogeneous pollen with high fertility and optimal morphometric parame

The needles and pollen of Pinus densiflora and Pinus koraiensis were studied for their lipid contents and fatty acid composition. The total lipid contents in needles of Pinus densiflora and Pinus koraiensis were 5.0 and 4.5%, whereas in pollen of Pinus densiflora and Pinus koraiensis 3.5 and 5.6%, respectively. Twenty-four fatty acids ranged from lauric acid to docosahexaenoic acid(22:6Ω3) were identified in the needle lipids. In needles, linolenic acid and palmitic acid were the major fatty acids. The needles of Pinus densiflora showed higher proportions of docosahexaenoic acid and 5-olefinic nonmethylene-interrupted polyenoic acids than those in the Pinus koraiensis. Twenty fatty acids ranged from myristic acid to lignoceric acid were identified in the pollen lipids. Linoleic acid was the major fatty acid in the pollen followed by oleic and palmitic acid. The fatty acid profile of pollen of Pinus densiflora was similar to those of the Pinus koraiensis pollen lipids.

This study was conducted to determine the optimum conditions for the treatment of polyamine to promote pollen germination and improve the fruiting rate by overcoming fertilization defects in low-temperature strawberry cultivation. The optimum temperature for pollen germination of strawberries was 30 for Seolhyang and 25 for Maehyang, and Seolhyang had a higher pollen germination rate than that of Maehyang. The addition of polyamines, namely spermine and putrescine to the medium improved the pollen germination rate and spermine treatment showed a better effect than putrescine treatment. The proper polyamine type and treatment concentration for enhancing the potency of pollen germination was 500 μM of spermine for both Seolhyang and Maehyang, which improved the pollen germination rate by 19-23% compared with that of the control. However, combined treatment of spermine and putrescine, resulted in a lower germination rate lower than that of the single treatment. Our results indicated that the treatment of polyamines during flowering in protected cultivation of strawberrise can improve the fruiting rate by overcoming the problem of poor pollen germination due to low temperature.

The purpose of this study is to clarify the environmental changes and human activities before and after the fortification of the Seongsan Sanseong Fortress (hereinafter mountain fortress) in Haman through using pollen analysis. For this purpose, samples were taken at the Tr-2 site in the Warm-temperate forest. The results of the analysis are as follow. ① Before and after the fortification of the mountain fortress, the area around it has changed from a broad-leaved forest (The HS-I period) into a deciduous broad-leaved forest (The HS-II period) and finally into a the mixed conifer and deciduous broad-leaved forest (The HS-III period). ② The HS-I period (before the middle of the 6th century, compared to the nationwide pollen zone RIIIa during the Postglacial) was covered with cool-temperate southern forests(including some species of Warm-temperate trees) and was very similar to the current climate environment during the Gaya period before the fortification. ③ The HS-II period (mid to late 6th century, pollen zone RIIIa) was covered with typical cool-temperate southern forests after the fortification and the temperature was slightly lower than the first stage. ④ The HS-III period (7th~8th century/9th century, pollen zone RIIIb) is an era of clear human interference, causing the destruction of forests in earnest and paving an active agricultural life.

Senna tora (L.) Roxb. belongs to Leguminosae and its seeds are usually roasted and boiled in water to produce tea in Korea. Also the plants are well known for the treatment of Hypertension, Hepatitis, Constipation and Conjunctivitis. This study was conducted to investigate the morphological characteristics and pollen germination rates of Senna tora with the aim of genetic mapping of this species. First, we investigated morphological characteristics of domestic and international genotypes containing 51 lines in Korea and 2 lines in China. No significant differences in growth characteristics were observed among 53 genotypes. However, ST-9 line which was collected in Pyung-chang showed high growth rate at the early stage. The flower of Senna tora consists of 5 petals, 10 surgeries (7 main surgeries, 3 small surgeries), 1 pistil, and 5 sepals. After bud emergence, each petal was white in 1-2 days, turn into ivory in a week, then yellow in 8-9 days and finally bloomed in 9-10 days. Although the average flowering times of the plants were July 24, ST9 flowered in July 12. In addition, the flowers of ST9 differed from the flowers of the other genotypes. Flower of ST9 joined together with one another. Therefore, ST9 showing high growth rate at the early stage and unique flowering characteristics was selected to as the paternal line for genetic mapping study. Second, we investigated pollen germination rates for each stage of flower development. Pollen started to germinate at the yellow bud stage and pollen germination rate was highest in the bloomed flower stage. This results show that self-fertilization hardly occurs when the flower is ivory bud stage, and there is no need to use young bud flowers for artificial crossing. This work is intended to serve as the basis for the breeding of new varieties in Senna tora.

Map-based cloning is a basic method for identifying the mutated gene in plants. We selected the gametophytic mutant, named as AP-26-09, in activation-tagging pool. Mutant plant showed various kinds of pollen phenotype, such as the different number of nucleus or abnormal shapes. For the map-based gene cloning, we conducted phenotypic analysis of F2 mapping population through the screening of DAPI-stained pollen using fluorescence microscopy. Genomic DNA of F2 plants is prepared from leaves of approximately 1000 plants. In order to define chromosomal region where mutation is located, we designed SSLP markers and performed PCR amplification. In this study, we characterized gametophytic mutant and determined the chromosomal location using map-based approach.

To identify genes that play critical roles during male gametogenesis in Arabidopsis, we have isolated several pollen morphological mutants from a mutagenized seed pool generated with a T-DNA activation vector. In this study, we have focused on a mutant plant producing ~50% abnormal pollen grains including high levels of collapsed pollen at maturity. The pollen developmental analysis showed that the mutant pollen phenotype was first observed at tricellular stage. Interestingly, the mutation was only maintained as a heterozygote due to the severely reduced genetic transmission through both sexes. TAIL PCR analysis led to the identification of the responsible gene which encodes a conserved oligomeric golgi complex component-related protein (COGCC). RT-PCR analysis showed predominant expression of the gene in reproductive organs including developing spores. The gene identity was confirmed by the result that mutant plants harboring a T-DNA containing corresponding wild type gene produced less level of mutant pollen grains. Furthermore, confocal laser scanning microscopy using mature pollen expressing COGCC-RFP driven under the native promoter showed small punctate signals, which are likely to be from the Golgi complex. Further experiments for co-localization of the COGCC-RFP with the Golgi markers are underway.

Tissue-specific promoters are a very useful tool for manipulating gene expression in a target tissue or organ; however, their range of applications in other plant species has not been determined, to date. In this study, we identified two late pollen-specific rice promoters (ProOsLPS10 and ProOsLPS11) via meta-anatomical expression analysis. We then investigated the expression of both promoters in transgenic rice (a homologous system) and Arabidopsis (a heterologous system) using ProOsLPS10 or ProOsLPS11::GFP-GUS constructs. As predicted by microarray data, both promoters triggered strong GUS expression during the late stages of pollen development in rice, with no GUS signals detected in the examined microspores and sporophytic tissues. Interestingly, these promoters exhibited different GUS expression patterns in Arabidopsis. While in Arabidopsis, the OsLPS10 promoter conferred GUS expression at the uni- and bi-cellular macrospore stages, as well as at the shoot apical region during the seedling stage, the OsLPS11 promoter was not active in the pollen at any stage, or in the examined sporophytic tissues. Furthermore, by performing a complementation analysis using a sidecar pollen (scp) mutant that displays developmental defects at the microspore stage, we found evidence that OsLPS10, which can be an applied promoter expressed in Arabidopsis, is useful for directing gene expression in the early stages of pollen development. Our results indicate that the OsLPS10 and OsLPS11 promoters can drive the expression of target genes during the late stages of pollen development in rice, but not in Arabidopsis. Our results also emphasize the necessity of confirming the applicability of an established promoter to heterologous systems.

부안지역 소나무 집단의 화분유동과 교배양식 모수를 추정 하기 위하여 7개 microsatellite 표지로 모수, 주변 성목 및 종자 에 대한 유전변이를 분석하였다. 이형접합도 기대치(He)와 근 교계수(F)는 각각 모수에서 0.614과 0.018, 종자에서 0.624과 0.087이며, 각 세대간에 차이는 없었다(P ＞ 0.05). MLTR로 추 정한 타가교배율(tm)은 0.967이며, 양친간 근연계수(tm-t s)는 0.057, 부계상관(rp)은 0.012로 나타났다. 기존에 보고된 소나 무의 동위효소 분석 결과에 비하여 타가교배율은 높고 근친교 배 및 부계상관은 낮았으나, microsatellite 표지를 이용한 소나 무류의 결과들과는 유사하였다. TwoGener로 추정한 최적 화분 비산 모델은 유효밀도(d = 220 trees/ha)를 가정한 정규확산모 델로 판명되었으며, 평균 화분비산거리(δ )는 11.42 m로 계산되 었다. 화분원 유전적 분화(Φft)는 0.021이며, Mental 검증에서 모수간 지리적 거리와 화분원의 유전적 분화는 상관성이 없는 것으로 나타났다(r = -0.141, P ＞ 0.05). 부안지역 소나무 집단 은 대부분의 화분이 가까운 거리에서 공급되지만, 화분수의 유 전다양성이 높고 화분원의 유전적 차이가 작은 상태로 추정된 다. 이러한 조건에서 완전한 임의교배가 이루어지기 때문에 종 자의 유전자형이 다양하며 세대간 유전변이의 감소가 없는 것 으로 사료된다.

The correct development of male gametophytes (pollen grains) in flowering plants is essential for proliferate in gamete production. Here we have taken a map-based cloning approach using Arabidopsis male gametophytic mutant, named gemini pollen3 (gem3) to identify and characterize key gene that is expressed gametophytically for the completion of microgametogenesis focusing on genes which control cell division and cell fate determination. Previously reported gem1 and gem2 mutants with similar characteristics to gem3 that are disturbed at asymmetric division and cytokinesis at pollen mitosis I (PMI) in Arabidopsis. However, gem3 was mapped to a different genetic locus, and pollen developmental analysis revealed that gem3 exert an effect at meiosis and mitosis causing complete sterility. We also discovered that gem3 homozygous lines produce aberrant pollen grains, arising from incomplete cytokinesis during male meiosis with sporophytic phenotypes of twisted-shape leaves, large flowers. This mutation shows reduced genetic transmission of gem3 allele through male gametophyte. In previous results, the gem3 locus was confirmed by mapping to the region located on chromosome 5. To further confirm strong candidate gene, we performed sequencing and genetic complementation analysis. Currently, we are performing functional studies of the gem3 gene for the better understanding of molecular mechanisms that control asymmetric division at meiosis and mitosis during pollen development.

Based on the results of microarray analysis we selected ten candidate genes that express in pollen at the early pollen developmental stage. By PCR amplification, the promoter region of these genes were amplified from rice genomic DNA (Nipponbare) and cloned into the destination pKGWFS7 vector via an entry vector, pDONR201. The characteristic of promoters were evaluated in Arabidopsis thaliana (Col-0) through GUS expression analysis. Fifty T2 plants respectively from each promoter were tested. Whole inflorescence of individual plant was stained with 1mM X-Gluc solution to observe tissue-specific GUS expression patterns. The results showed that all 10 promoters activated in pollen tissues. Among them six promoters expressed at the early developmental stage (unicellular) of pollen and the others expressed at both early (unicellular) and late pollen developmental stage (mature pollen). The results indicated that these promoters would be potential applicable for the studies of pollen function. Currently, we are performing these promoters analysis in rice transgenic plants as well as molecular characterization.

Wheat is one of the most important food crops in the world and provides 20% of nutritional requirement to the consumers in developing countries. The consumption per capita of wheat is persistent but the self-sufficiency has decreased owing to the less competitive world market and double cropping system with rice in Korea. This study was conducted to obtain baseline data on the selection lines with early maturity and good agronomic traits. Geumgangmil, a leading Korean variety, was crossed with CIMMYT’s line, and a DH (double haploid) line was released using pollens of Maize in Mexico. The 71 inbred lines were grown in fields at NICS, Miryang. The culm length of inbred lines were normally distributed from 37cm to 106cm, though that of the P1 (Geumgangmil) was 71 and P2 (CIMMYT’s Line) was 77cm. Especially, some lines were very short and measured below 50cm. The spike lengths of most of the lines were longer than that of P1 (7.5cm), but were normally distributed. The number of spikelet was less than that in P2 (19ea.), but was more than P1(10ea.). The heading day of most of the lines was between P1 (11th April) and P2 (25th April) and the maturity day of the lines was similar to P1 (1st June) but 5 lines showed 2~6 days shorter maturity period than that of P1. However, 20 lines were found to have 2~10 days longer maturity period than that of P2.

This study was carried out to investigate the effect of sucrose, boric acid, and calcium nitrate in optimum pH and relative humidity on pollen germination and pollen tube growth of boxthorn (Lycium chinense M.). Medium containing 100mg/L of H3BO3 showed maximum pollen germination and pollen tube elongation, 64.8% and 920μm. Additionally supplied calcium nitrate to the medium lowered pollen germination rate and pollen tube elongation. Here we report the optimum conditions for the pollen germination and pollen tube development was observed at 10% sucrose, 1.0% agar, pH 6.0, and 100% relative humidity.