벼는 세계 인구의 절반이상의 안정적 식량으로 가장 중요한 식량자원이라고 할 수 있으며, 미래의 품종육종을 위해 사라져가는 재래종 자원과 벼생산에 영향을 미칠 생물적, 무생물적 스트레스에 조화를 이룰 수 있는 근연야생종을 보존하고자 1977년 국제미작연구소내에 종자은행을 설립하였다. 현재 123개국으로부터 109,000여점의 유전자원을 확보하여 보존하고 있으며, 야생종 자원도 5,000여점 포함되어있다. T.T. Chang GRC에 새로 도입된 종자는 IRRI내의 SHU(Seed Health Unit)에서 검역을 마친 후 증식을 수행할 수 있다. 증식은 주로 건기에 실시하며 생태형, 원산지, 광감수성 유무 등을 고려하여 파종기와 수확기를 달리함으로서 충실한 종자를 확보하는데 주력하고 있다. 증식된 벼 종자는 건조를 거쳐 active collection 과 base collection 으로 나누어 보존하며, 안전중복보존을 위하여 제3국인 스발바르국제저장고와 미국의 국립유전자원보존센터에도 보존하고 있다. 보존자원의 종자량, 발아율 모니터링을 통하여 갱신을 결정한다. 보존된 자원은 연구목적으로 분양 및 교환을 통하여 활용된다. 유전자원의 정보관리는 GRIMS(Genetic Resources Information Management System)로 운영되고 있다.

This study describes an efficient and stable droplet vitrification following cryopreservation of strawberry shoot tip (Fragaria × ananassa Duch.) accessions ‘Massey’ and ‘MDUS3816’. The shoot tips were precultured in Murashige and Skoog (MS) liquid medium supplemented with sucrose (0.3-0.7M). Precultured explants were osmoprotected with loading solution (LS, C4) containing 17.5% glycerol and 17.5% sucrose for 40 min and exposed to dehydration solution (B1) containing 50% glycerol and 50% sucrose for 40 min at 25oC. Subsequently, the explants were transferred onto droplets containing 2.5 μL PVS3 on sterilized aluminum foils (4 ㎝× 0.5 ㎝) prior to direct immersion in liquid nitrogen (LN) for 1 h. The highest regrowth rate (%) in both the cultivars was obtained when the shoot tips were precultured with 0.3M sucrose for 30 h + 0.5M sucrose for 16 h at 25oC. The cryopreserved shoots tips exhibited 57.8 % recovery rate by culturing in NH4NO3-free MS medium supplemented with 3% sucrose, 1.0 g/L casein, 1.0㎎/L GA3, and 0.5 ㎎/L BA for 5 weeks and in MS medium supplemented with 0.5 ㎎/L GA3 for 8 weeks. Variation was not observed in both of ploidy analysis and morphological investigation on plantlets of two accessions cryopreserved under variable preculture conditions.

This study describes an efficient and widely applicable droplet-vitrification following cryopreservation for shoot tips of strawberry (Fragaria × ananassa Duch.) cvs. ‘Wonkyo3114’ and ‘Gurumi40’. The shoot tips were precultured in Murashige and Skoog (MS) liquid medium supplemented with sucrose (0.3-0.5M). Precultured explants were osmoprotected with loading solution (LS, C4) containing 20% glycerol and 20% sucrose for 40 min and exposed to dehydration solution (B5) containing 40% glycerol and 40% sucrose for 40 min at 25℃, Subsequently, the explants were transferred onto droplets containing 2.5 μL PVS3 on sterilized aluminum foils (4 ㎝ × 0.5 ㎝) prior to direct immersion in liquid nitrogen (LN) for 1 h. The highest regrowth rate (%) in both the cultivars was obtained when the shoot tips were precultured with MS + 0.3M sucrose for 40 h at 25℃. The cryopreserved shoots tips exhibited 55% regrowth rate by culturing in NH4NO3-free MS medium supplemented with 3% sucrose, 1.0 g/L casein, 1.0㎎/L GA3, and 0.5 ㎎/L BA for 5 weeks and in MS medium supplemented with 0.5 ㎎/L GA3 for 8 weeks. This result shows that droplet-vitrification could be employed as a promising method for cryostorage of strawberry germplasm.

This study describes the successful establishment of a cryopreservation protocol for Citrus limon cultivars: ‘Frost Eureka limon’ and ‘Cook Eureka limon’, using a droplet-vitrification method. The shoot tips that were excised from in vitro grown seedlings of the two cultivars were preserved in liquid nitrogen (LN) and successfully regenerated into whole plants. Excised shoot tips were pre-cultured for 1 or 2 days in 0.3 M and 0.5 M sucrose solutions at 25℃ and incubated in a loading solution (LS) composed of 17.5% glycerol + 17.5% sucrose in Murashige and Skoog (MS) medium for 40 min at 25℃. Prior to direct immersion in LN for 1 h, the shoot tips were dehydrated with plant vitrification solution 2 (PVS2) at 0℃ or PVS3 at 25℃. The frozen shoot tips were re-warmed and unloaded with 1.2 M sucrose in ½ MS for 30 min at 25℃. Shoot tips were post-cultured overnight on survival medium and then micrografted onto ‘trifoliate orange’ (Poncirus trifoliate (L.) Raf. seedling rootstocks for recovery and to produce whole plants. The highest regrowth rates were 53.5% and 50.3% for cryopreserved shoot tips of ‘Frost Eureka limon’ and ‘Cook Eureka limon’, respectively, when pre-cultured in 0.3 M and 0.5 M sucrose concentrations in a sequencing manner, with LS and treated with PVS2 for 60 min at 0℃. We also investigated whether the ammonium ion concentration on post-culture medium affected the viability of the cryopreserved Citrus shoot tips. The viability of cooled samples, following culturing on woody plant media (WPM) containing ¼ ammonium nitrate overnight before micrografting, was the highest (70.3%) in ‘Frost Eureka limon’. The study described here is a cost-effective and safe method to conserve Citrus fruit cultivars, for the improvement and large-scale multiplication of fruit plants and for breeding disease resistance.

This study describes an efficient and widely applicable droplet-vitrification following cryopreservation for shoot tips of Chrysanthemum morifolium (Ramat.) cvs. ‘Borami’ and ‘Yes morning’. The shoot tips of Chrysanthemum were precultured in Murashige and Skoog (MS) liquid medium supplemented with sucrose (0.3-0.7 M). Precultured explants were treated with loading solution (LS, C6) containing glycerol 20% and sucrose 20% for 30 min and exposed to dehydration solution (B5) containing 40% of glycerol and 40% of sucrose for 60 min at 25℃, and then transferred onto droplets containing 2.5 ㎕ PVS3 on sterilized aluminum foils (4 ㎝ × 0.5 ㎝) prior to direct immersion in liquid nitrogen (LN) for 1 h. The highest regeneration rate (%) was obtained when shoot tips were precultured with treatment-2 (exposing of shoot tips to MS + 0.3M sucrose for 30 h and then treated with MS+0.5 M sucrose for 16 h) at 25℃ in both the cultivars. The viability of cooled samples, followed by culturing on NH4NO3-free MS medium for first 5 days was increased to two-fold (80.7%) regrowth rate over those cultured on normal MS medium or MS medium containing plant growth regulators. This result shows droplet-vitrification would be a promising method for cryobanking chrysanthemum germplasm.

Plant regeneration via organogenesis and somatic embryogenesis was investigated in Korean soybean cultivars including Cheongja 3, Jinpumkong 2, Taekwangkong and Uram. Cotyledon, cotyledon+hypocotyl and hypocotyl segments of 7-day-old seedlings were cultured on MS medium containing various concentration (0, 1, 2 and 4 ㎎/L) of BA and TDZ. The results showed that MS medium supplemented with BA 2.0 ㎎/L yielded the highest shoot formation ratio of 83.3%. In 4 cultivars, Taekwangkong showed the highest ratio of shoot formation. When various sizes of immature cotyledons (S: 1∼ 2 ㎜, M: 3∼5 ㎜, L: 6∼8 ㎜) were tested on MS medium containing 2,4-D 40 ㎎/L for somatic embryogenesis, the optimum size for embryogenic callus induction was 3∼5 ㎜ in length of immature cotyledons. In 4 cultivars, Taekwangkong showed the highest percentage of embryogenic callus induction. The results indicate that Taekwangkong is the best soybean cultivar for plant regeneration via organogenesis and embryogenic callus induction among the 4 cultivars.

The genus Allium which is one of the biggest plant genera comprises around 750 species that show the various morphological and ecological diversity with various taxonomic and geographical groups. The species of genus Allium has not been clearly distinguished because of extraordinary large amount of variation. We developed 8 simple sequence repeat markers (SSRs) which showed polymorphism within A. sativum accessions, but it was insufficient in transferability analysis of other Allium species. In this regard, we finally selected 50 primer pairs which was applicable to other Allium species adding to 8 primer pairs. According to results from application of 50 SSR markers to 9 Allium species, the average number of alleles ranged from 1.400 (A. porrum) to 1.889 (A. tuberrosum) and A. tuberosum (2n=32) has maximum 9 alleles. The lowest transferability value was 42.0% ( A. cepa and A. chinense) while A. sativum showed 96.7%. The A. porrum conceived to be close relationship to A. sativum as Allium subgenera revealed higher transferability (73%) rather than other Allium species. According to PCA analysis, three groups were divergent, and the A. fistulosum and A. sativum revealed the distinct groups and the rest 7 species formed another group.

Identification of compatible parental line is of great importance in introduction of useful characters to onion breeding program, beyond the severe hybridization barrier. Phylogenic analysis of Allium section Cepa was conducted through PCR by URPs, repeated sequences of A. fistulosum, and microsatellite markers. Totally 76 accessions originated from 21 countries were clustered into five groups at a 0.84-similarity level: group I;A. cepa and its wild relatives and A. cepa ssp. ascalonicum, group II; A. cepa ssp. wakegii, A. cepa ssp. proliferum and Samcheung-pa group III; A. fistulosum and A. altaicum, group IV; A. galanthum, group V; Soeckkori-pa. Samcheung-pa and Soekkori-pa, Korean local varieties, shared band type of both Cepa group and Altaicum group, indicating that those are derived from interspecific hybridization between A. fistulosum and A. cepa.

Allium is one of the largest genera, which has more than 700 species. PCR by URP (universal rice primer) primers was carried out to get phylogenetic information on 26 species, 62 accessions of subgenus Rhizirideum. The accessions were divided into seven groups at 0.76 similarity level. A. tuberosum (Chinese chives) and A. ramosum represented high similarity of 0.91. A. montanum, A. nutans, A. senescens, A. libani, A. odorum, A. austrosibiricum, and A. narcissiflorium grouped at 0.80 similarity. Some of the wild species, such as A. prostratum, A. polyrhizum, A. odorum, and A. mongolicum, showed different band patterns according to polyploidy, occurrence of B-chromosome, collection site, and origin.