버즈풋 트레포일 (Lotus corniculatus L. cv. Empire) 종자로부터 직접 캘러스를 유도하고, 형성된 캘러스로부터 식물체를 재분화하는 조건을 확립하였다. 캘러스 유도시 사용한 SH (Schenk and Hildebrandt), MS (Murashing and Skoog), N6 (Chu) 배지중에서 SH 배지가 캘러스 유도에 유리한 것으로 나타났으며, 캘러스 유도 및 중식시에는 2,4-D 3 m g / ℓ 을 첨가한 조건이 가장 효율이 좋았다. 식물체 재분화 조건은 BOi2Y 배지에서 20일 간격으로 계대배양하며 재분화를 완성한 조건 2가 더 좋았으며, 캘러스로부터 완전한 식물체로 재분화되는데 필요한 시간은 약 60일이었다.

목초의 내하고성을 증진시킬 목적으로 이용하고자 하는 내열성 유전자 (BcHSP17.6)의 발현양상을 배추에서 조사하였다. BcHSP17.6 단백질을 항원으로 한 항체를 생산하여 항원-항체 반응으로 확인한 결과, 생산된 항체는 항원과 결합하므로서 항체가 정상적으로 생산되었음을 확인하였다. 이 항체를 이용하여 다양한 heat shock (HS) 조건에서 15-~18-kD low molecular weight heat shock proteins (LMW HSPs)가 축적되는지를 조사하였다. 이들 LMW HSPs는 $35^{\circ}C$ 수식 이미지 처리에서 나타나기 시작하였으며, $40^{\circ}C$ 수식 이미지에서 4시간 처리하였을 때 total protein 100 mg당 $1.56{\mu}g$ 수식 이미지이 축적되어 최대치를 나타내었다. 또한 합성된 이들 LMW HSPs는 HS 처리후 24시간이 경과되어도 거의 변함이 없었으며, $40^{\circ}C$ 수식 이미지 보다 높은 온도 조건에서는 축적량이 감소하였다. 이와 같이 일단 합성된 LMW HSPs가 장시간 동안 감소하지 않는다는 사실로부터, 이들 HSPs들이 식물체가 내열성을 가지도록 하는데 관여하며, 더 나아가서 배추가 정상 생육온도보다 더 높은 치사온도에서도 살아남을 수 있도록 하는데 중요한 역할을 하는 것으로 사료된다.

IT기술 진보로 인하여 국내 복화운송(Street Turn) 시스템도 많은 발전을 거듭하고 있지만 복화운송 물량의 증가로 귀결되지 않고 있다. 이러한 측면에서 본 연구는 기존 복화운송 시스템을 사용하고 있는 사용자 입장에서 시스템을 이용하게 하는 의사결정 요인을 제시하는데 연구의 목적이 있다. 퍼지계층분석(Fuzzy-AHP)을 이용하여 국내 복화운송을 사용한 경험이 있거나 해운기업에 종사하고 있 는 경력자를 대상으로 분석을 실시하였다. 총 5개의 대요인과 17개 세부 요인을 추출하였으며, 연구결과 대요인으로 적시성 요인이 가장 중요한 요인이며 그중 정보제공 시기(0.207)가 가장 중요한 요인으로 선정되었으며, 다음으로 플렛폼 이용 프로세스(0.079), 참여 화주의 수 (0.074)가 뒤를 이었다.

Small heat shock proteins (Hsps) are one of most conserved molecular chaperones that protect stress-inducible denaturation of substrates in living organisms. Small Hsps consist of a large subfamily categorized by subcellular localization ranging in size from 12 to 40 kDa. Here, we identified and characterized a small Hsp 16.9 gene (EsHsp16.9) from Siberian wild rye (Elymus sibiricus L.). EsHsp16.9 is a 456-bp cDNA with an open reading frame predicted to encode a 151-amino acid protein. It possesses a conserved ɑ-crystallin domain, which is a unique domain for small Hsps; shares high sequence similarity with cytosolic class I small Hsps among the small Hsp subfamily in Arabidopsis; and is close (96% similarity) to small Hsp in wheat. Northern blot analysis showed that EsHsp16.9 transcripts were enhanced by heat, drought, arsenate, methyl viologen, and H2O2 treatments. Moreover, we expressed and purified recombinant EsHsp16.9 proteins in Escherichia coli to confirm its activity as a molecular chaperone. We found that recombinant EsHsp16.9 exhibits effective molecular chaperone activity, as determined by inhibition of thermal aggregation of malate dehydrogenase (MDH), which is broadly used as a model substrate.

To develop transgenic forage crops with enhanced tolerance to abiotic stress, we introduced an alfalfa Hsp23 gene expression vector construct through Agrobacterium-mediated transformation. Integration and expression of the transgene were confirmed by PCR, northern blot, and western blot analyses. Under normal growth conditions, there was no significant difference in the growth of the transgenic plants and the non-transgenic controls. However, when exposed to various stresses such as salt or arsenic, transgenic plants showed a significantly lower accumulation of hydrogen peroxide and thiobarbituric acid reactive substances than control plants. The reduced accumulation of thiobarbituric acid reactive substances indicates that the transgenic plants possessed a more efficient reactive oxygen species-scavenging system. We speculate that the high levels of MsHsp23 proteins in the transgenic plants protect leaves from oxidative damage through chaperon and antioxidant activities. These results suggest that MsHsp23 confers abiotic stress tolerance in transgenic forage crops and may be useful in developing stress tolerance in other crops.

In order to improve high persistence and forage quality, through selection of various superior parental varieties for breeding and synthesis of them with new lines, there are ongoing worldwide studies aiming to enhance the quality of tall fescue through a traditional breeding method by selection and hybridization. A new tall fescue variety (Festuca arundinacea Schreb.), named Greenmaster3ho, was developed by the National Institute of Animal Science, Rural Development Administration in Korea from 2010 to 2014. For synthetic seed production of this new variety, five superior clones, 09XFa02, 09XFa03, 09XFa11, 09XFa13, and 09XFa14 were selected and polycrossed. The agronomic growth characteristics and forage production capability of the seeds were studied at Cheonan from 2010, and regional trials were conducted in Cheonan, Hoengseong, Jeju, and Jinju from 2012 to 2014. Greenmaster3ho showed enhanced disease resistance, persistence, and regrowth ability as compared to Fawn. The dry matter yield of Greenmaster 2 was 29% higher (15,119 kg/ha) than that of Fawn. However, the nutritive value of both varieties was similar. This study developed a new tall fescue variety with excellent environmental adaptability, aiming to make a contribution to the vitalization of the Korean grassland industry.

Abiotic stress is the major limiting factor of forage crops growth and yields. The objective of this work was to study the stress tolerance and regeneration capability of transgenic forage crops carrying a MsHSP23 gene, encoding a alfalfa mitochondrial sHSP protein. The expression of the MsHSP23 gene was confirmed in bacteria, recombinant mHSP23 conferred tolerance to salinity and arsenic stress. Furthermore, mHSP23 was cloned in a plant expressing vector and transformed into forage crops such as alfalfa, tall fescue and bent grass. The transgenic plants exhibited enhanced tolerance to salinity and arsenic stress conditions. In comparison to wild type plants, transgenic plants were exhibited significantly lower electrolyte leakage. Moreover, the transgenic plants had superior germination rates when placed on medium containing arsenic. Taken together, these overexpression results imply that mHSP23 plays an important role in salinity and arsenic stress tolerance in transgenic forage crops. This approach could be useful to develop stress-tolerant plants including forage crops.

The molecular responses to various abiotic stresses were investigated by the approaches with transcriptomic analysis based on an ACP system. Here we identified differentially expressed genes under abiotic stresses in alfalfa seedlings and they were mostly unknown genes and a few common stress-related genes. Among them, mitochondrial small HSP23 was responded by the diverse stress treatment such as heat, salt, As stresses and thus it could be a strong candidate that may confer the abiotic stress tolerance to plants. When expressed in bacteria, recombinant MsHSP23 conferred tolerance to salinity and arsenic stress. Furthermore, MsHSP23 was cloned in a plant expressing vector and transformed into tobacco, a eukaryotic model organism. The transgenic plants exhibited enhanced tolerance to salinity and arsenic stress under ex vitro conditions. In comparison to wild type plants, the transgenic plants exhibited significantly lower electrolyte leakage. Moreover, the transgenic plants had superior germination rates when placed on medium containing arsenic. Taken together, these overexpression results imply that MsHSP23 plays an important role in salinity and arsenic stress tolerance in transgenic tobacco. The results of the present study show that overexpression of alfalfa mitochondrial MsHSP23 in both eukaryotic and prokaryotic model systems confers enhanced tolerance to salt and arsenic stress. This indicates that MsHSP23 could be used potentially for the development of stress tolerant transgenic crops, such as forages.

Tall fescue (Festuca arundinacea Schreb.) is an important cool season forage plant that is not well suited to extreme heat, salts, or heavy metals. To develop transgenic tall fescue plants with enhanced tolerance to abiotic stress, we introduced a MsHsp23 gene expression vector construct through Agrobacterium-mediated transformation. Integration and expression of the transgene were confirmed by PCR, northern blot, and western blot analyses. Under normal growth conditions, there was no significant difference in the growth of the transgenic plants and the non-transgenic controls. However, when exposed to various stresses such as salt or arsenic, transgenic plants showed a significantly lower accumulation of hydrogen peroxide and thiobarbituric acid reactive substances than control plants. We speculate that the high levels of MsHsp23 proteins in the transgenic plants protect leaves from oxidative damage through chaperon and antioxidant activities. These results suggest that MsHsp23 confers abiotic stress tolerance in transgenic tall fescue and may be useful in developing stress tolerance in other crops. Compared with traditional plant breeding, genetic engineering provides a relatively fast and precise means of achieving improved stress tolerance of forage crops. Development of forage crops that are more tolerant to various abiotic stresses could lead to the use of more new lands for cultivation.

Most forage crops growing under field conditions are often being exposed to various environmental stresses such as drought, freezing, high temperature, waterlogging and climate change. A combination of grass breeding approaches will likely be needed to improve significantly the environmental stresses tolerance of forage crops in the field. Attempts have been taken by grass breeders to develop tolerant varieties of different crops for environmental stress. A new tall fescue variety (Festuca arundinacea Schreb.) named ‘Purumi’ was developed by the National Institute of Animal Science, Rural Development Administration from 1999 to 2007. For synthetic seed production of this new variety, 5 superior clones, EFa9108, EFa0010, EFa0020, EFa0108, and EFa0202 were selected and polycrossed. The agronomic growth characteristics and forage production capability of the seeds were studied at Cheonan from 2004 to 2005, and regional trials were conducted in Cheonan, Pyungchang, Jeju, and Jinju from 2008 to 2010. ‘Purumi’ showed enhanced winter hardiness, disease resistance, and regrowth ability as compared to ‘Fawn’. The dry matter yield of ‘Purumi’ was about 5.6% higher as 16,821kg/ha than that of ‘Fawn’. However, the nutritive value of both varieties was similar. When this new variety of tall fescue, Purumi, has been developed and distributed with its most remarkable adaptability for Korean climates and superior value as a livestock feed, it is expected to play an important role for a new restoration of the pasture industry in Korea.

Italian ryegrass (Lolium multiflorum Lam.) is one of important forage crop grass widely cultivated in Korea. Progress in breeding using conventional selection procedure is very slow, since Italian ryegrass is highly self-infertile. Biotechnological approaches, therefore, may contribute to the development of improved cultivars for forage crops. In an effort to optimize tissue culture responses of Italian ryegrass (Lolium multiflorum Lam.) for future genetic manipulations to improve forage characteristics, the effects of culture medium supplements on tissue culture responses were investigated with mature seeds of Korean Italian ryegrass (Lolium multiflorum Lam.) seven cultivars as explant tissues.

Italian ryegrass (Lolium multiflorum Lam.) is one of important forage crop grass widely cultivated in Korea. The genetic manipulation of Italian ryegrass (Lolium multiflorum Lam.) necessitates a reliable and efficient, genotype-independent method of transformation. We are interested in developing molecular breeding methods to improve its nutritional quality and abiotic stress resistance. Development of a rapid and efficient transformation system is the basis for genetic manipulation of Italian ryegrass. In order to establish an efficient Agrobacterium-mediated transformation system was applied to transfer genes into seven genotypes of Italian ryegress, namely cv. 'Kogreen', 'Kopeed', 'Kowinearly', 'Kowinmaster', 'Hwasan 101', 'Hwasan104' and 'Kowinner.' The transformation system developed in this study would be useful for Agrobacterium-mediated genetic transformation of Italian ryegrass plants with genes of agronomic importance.