• Journal of Plant Biotechnology
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• v.44 no.1
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• pp.89-96
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• 2017

The CRISPR/Cas9 is a core technology that can result in a paradigm for breeding new varieties. This study describes in detail the sgRNA design, vector construction, and the development of a transgenic plant and its molecular analysis, and demonstrates how gene editing technology through the CRISPR/Cas9 system can be applied easily and accurately. CRISPR/Cas9 facilitates targeted gene editing through RNA-guided DNA cleavage, followed by cellular DNA repair mechanisms that introduce sequence changes at the site of cleavage. It also allows the generation of heritable-targeted gene mutations and corrections. Here, we present detailed procedures involved in the CRISPR/Cas9 system to acquire faster, easier and more cost-efficient gene edited transgenic rice. The protocol described here establishes the strategies and steps for the selection of targets, design of sgRNA, vector construction, and analysis of the transgenic lines. The same principles can be used to customize the versatile CRISPR/Cas9 system, for application to other plant species.

• Research in Plant Disease
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• v.21 no.2
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• pp.74-81
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• 2015

The specific and sensitive nested-PCR method to detect Acidovorax citrulli, a causal agent of bacterial fruit blotch on cucurbitaceae, was developed. PCR primers were designed from the draft genome sequence which was obtained with the Next Generation Sequencing of A. citrulli KACC10651, and the nested-PCR primer set (Ac-ORF 21F/Ac-ORF 21R) were selected by checking of specificity to A. citrulli with PCR assays. The selected nested-PCR primer amplified the 140 bp DNA only from A. citrulli strains, and detection sensitivity of the nested PCR increased 10,000 times of $1^{st}$ PCR detection limit (10 ng genomic DNA/PCR). The nested PCR detected A. citrulli from the all samples of seed surface wash (external seed detection) of the artificially inoculated watermelon seeds with $10^1cfu/ml$ and above population of A. citrulli while the nested PCR could not detected A. citrulli from the mashed seed suspension (internal seed detection) of the all artificially inoculated watermelon seeds. When the naturally infested watermelon seeds (10% seed infested rate with grow-out test) used, the nested PCR detected A. citrulli from 2 seed samples out of 10 replication samples externally and 5 seed samples out of 10 replication samples internally. We believe that the nested-PCR developed in this study will be useful method to detect A. citrulli from the Cucurbitaceae seeds.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.65 no.4
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• pp.496-507
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• 2020

Virus infections of apples result in lowered commercial qualities such as low sugar content, weakened tree vigor, and malformed fruits. An effective way to control viruses is to produce virus-free plants based on the development of an accurate and sensitive diagnostic method. In this study, real-time PCR assays based on SYBR Green I and TaqMan probes were developed for detecting ASGV, ASPV, and ApMV viruses. These methods can detect and quantify 10³ to 10¹¹ RNA copies/μL of each virus separately. Compared with methods with two different dyes, the SYBR Green I-based method was efficient for virus detection as well as for assay using the TaqMan probe. Field tests demonstrated that real-time PCR methods developed in this study were applicable to high-throughput diagnoses for virus research and plant quarantine.

• Smart Media Journal
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• v.7 no.3
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• pp.51-56
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• 2018

Banana is the eighth most important crop in the world designated by the United Nations Food and Agriculture Organization (FAO). In terms of production, it is the fourth most harvested crop following wheat, rice, and corn, considered to be a promising food source for mankind as it is not only scrumptious but also rich in nutrients. However, abnormal cultivation methods that mankind has invented to breed bananas is currently pushing them to the state of extinction. The aim of this study is to develop information-rich digital contents that can easily and intriguingly convey information about steadily but certainly increasing risk of banana extinction. The end-users are middle and high school students who are assumed to be aware of the agricultural revolution through their school curriculum. The linear storytelling method was meant to encourage the end-users to be interested in the causes and processes in which banana has become endangered. For generating a natural interaction between the content and the end-users, a parallel, upper-lower structure was used, in which the relationship between the stories comprising several stages and the information graphic supporting each story is represented. This paper can support an information campaign about the endangered species.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2020.12a
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• pp.56-56
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• 2020

감초는 다년생 콩과(Leguminocae) 식물로 국내에서 시중가격이 높은 만주감초가 일부 재배되고 있다. 우리나라에서 감초 재배법이 불완전한 상황에서 한반도의 기후변화에 의한 온도 상승은 약용작물의 생산 및 품질에 많은 영향을 미칠 것으로 예상되므로 본 연구에서는 재배환경 중 온도 조건만 조절할 수 있는 온도구배터널(temperature gradient tunnel system)을 이용하여 4개의 T1(외기온도+0.5~1.3℃), T2(+1.3~2.2℃), T3(+2.2~3.2℃), T4(+3.2~4.0℃) 처리로 온도구배 하여 4년생 만주감초(Glycyrrhiza uralensis F.)를 재배하였다. 지하부가 오래된 모주와 신초1의 경우 저온(T1)과 중간구간(T2, T3)에서 초장과 총화수가 우세하였고, 번식이 가장 늦은 신초2의 경우 중간구간(T2, T3)에서의 생육 및 개화반응이 뚜렷했다. 각 온도처리구마다 3개의 감초 개체를 선발하여 모주의 정단으로부터 5개의 성엽을 채취하였다. Reverse transcription quantitative PCR (RT-qPCR)은 AccuPower® GreenStar^TM RT-qPCR Master Mix (Bioneer, Korea)를 이용하여 진행되었다. Primer 디자인은 NCBI Primer-blast 프로그램을 사용해 제작하였고 ABI StepOne real time system (Applied Biosystem)의 melting curve analysis에서 one-peak test를 통해 gene specific primer임을 확인하였다. 각 온도처리구의 감초 잎에서 RNA를 추출하였고, RT-qPCR을 통해 감초의 유전자 발현양상을 비교, 분석하였다. Phytochrome interacting factor 4 (PIF4)는 식물 호르몬을 유발하는 전사조절을 조정함으로써 고온 신호전달에 핵심적인 역할을 수행한다. 활성화된 Phytochrome B(PhyB)는 PIF4의 활성을 억제한다고 알려졌다. Eukaryotic initiation factors(eIFs)는 mRNA 번역 개시인자로 유전자 발현과 특정 단백질 생산을 조절하는 역할을 한다. 본 결과는 온도조건에서 반응하는 생리적 변화를 전사체 수준에서 조사 분석하여 생리해석의 기초자료로 활용, 국내 감초 재배를 위한 환경조건 구명 및 적지 선정 기초자료로서 활용을 기대한다.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.3
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• pp.41-48
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• 1965

This study was aimed at knowing the magnitude of year variation in rice heading dates under the different seasonal cultures, and to estimate the heading date in advance. Using six rice varieties such as Kwansan, Suwon＃82, Suwon ＃144, Norin＃17, Yukoo＃132 and Paltal, the early, ordinary and late seasonal cultures had been carried out at Paddy Crop Division, Crop Experiment Station at Suwon for the six-year period 1959 to 1964. In addition the data of the standard rice cultures at the Provincial Offices of Rural Development for the 12-year period 1953 to 1954, were analyzed for the purpose of clarifying a relationship between variation of rice heading dates and some of meteorological data related to the locations and years. The results of this study are as follows: 1. Year variation of rice heading dates was as high as 14 to 21 days in the early seasonal culture and 7 to 14 days in the ordinary seasonal culture, while as low as one to seven days in the late seasonal culture which was the lowest among three cultures. The magnitude of variation depended greatly on variety, cultural season and location. 2. It was found out that there was a close negative correlation between the accumulated average air temperature for 40 days from 31 days after seeding and number of days to heading in the early seasonal culture. Accordingly, it was considered possible to predict the rice heading date through calculation of the accumulated average air temperature for the above period and then the linear regression(Y=a+bx). On the other hand, an estimation of the heading date in the late seasonal culture requires for the further studies. In the ordinary seasonal culture, no significant correlation between the accumulated average air temperature and number of days to heading was obtained in the six-year experiments conducted at Suwon. There was a varietal difference in relationship between the accumulated average air temperature for 70 days from seeding and number of days to heading in the standard cultures at the provincial offices of rural development. Some of varieties showed a significant correlation between two factors while the others didn't show any significant correlation. However, there was no regional difference in this relationship.

• Proceedings of the Korean Society of Postharvest Science and Technology of Agricultural Products Conference
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• 2003.04a
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• pp.139-139
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• 2003

원예작물은 수확 후 여러 가지 요인에 의해 품질이 저하되어 상품성을 잃게되는데, 특히 수확 후 품온상승에 의해 호흡작용이 왕성해져 품질변화를 촉진하게 된다. 그러므로 수확 후 품온을 가능한 한 빠르게 낮추어 신선도를 유지하기 위한 예냉, 저온저장, 저온수송기술등 저온유통시스템을 활용하는 것이 매우 필요하다. 저온유통시스템을 효과적으로 활용하기 위해서는 냉기의 순환이 원활하여 냉각효율이 좋고 파손의 위험과 물류비용을 줄일 수 있는 포장상자의 구조에 대한 연구가 필요하다. 포도, 방울토마토, 참외의 소비형태는 핵가족화와 더불어 신선 고품질의 농산물을 소량 구매하는 소비패턴으로 변화하면서 저온유통 및 소포장을 위한 포장형태의 개선이 요구되고 있다. 따라서 예냉.저온유통에 적합한 포장상자를 개발하기 위하여 연구소, 농협중앙회, 상자제작회사, 포장디자인 업체, 산지농협 작목반이 개발협의체를 구성하여 참외, 포도, 방울토마토의 저온유통용 골판지 상자를 개발하여 압축강도 및 냉각시험을 실시하여 소비지에 시범 보급하였다. 1. 참외 포장상자는 3kg용 상부 개방형 골판지상자로써 손잡이를 부착하고, 포장상자 상부에 덮개용 필름을 부착하였다. 제작된 상자의 압축강도는 저온유통 후 에도 안전압축강도 이상인 343kgf의 압축강도를 나타냈고, 냉각효율도 기존상자에 비하여 높게 나타났다. 또한 개발된 상자를 사용하여 시중에 출하할 경우 기존 5kg 상자에 비하여 8%이상의 부가가치를 높일 수 있었다. 2. 포도 포장상자는 소비자 기호도를 충족하고 다층적재시 손상을 방지하기 위하여 내용물을 1단 적재하고 상자를 개방형으로 하여 덮개용 필름을 부착하므로써 소비자가 내용물을 확인할 수 있도록 하고, 상자의 압축강도는 소비지까지 유통이 완료된 후 에도 수직압축강도가 400kgf이상으로 안전압축강도 보다 높아 저온에서 안전하게 유통할 수 있었다. 또한 개발된 상자를 사용하여 시중에 출하할 경우 기존 5kg 상자에 비하여 2kg상자는 21%, 4kg상자는 12%이상의 부가가치를 높일 수 있었음. 3. 방울토마토 포장상자는 외포장 상자를 4kg용 상부 개방형 골판지상자를 사용하고, 내포장상자는 500g용 PET용기를 사용하여 1상자에 8개의 내포장상자가 적재될 수 있도록 제작하였다. 제작된 외포장상자의 압축강도는 저온유통후에도 320kgf으로 안전압축강도보다 높게 나타났고, 파랫트 적재효율도 96%이상으로 나타나 물류의 효율성을 높일 수 있었다. 개발된 상자를 사용하여 시중에 출하할 경우 기존 산물형태의 4kg용 상자에 비하여 500g 내포장상자에 출하할 경우 20%이상의 부가가치를 높일 수 있었다.

• Journal of Plant Biotechnology
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• v.48 no.1
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• pp.34-43
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• 2021

Targeted genome editing using the CRISPR/Cas9 system is a ground-breaking technology that is being widely used to produce plants with useful traits. However, for woody plants, only a few successful attempts have been reported. These successes have used Agrobacterium-mediated transformation, which has been reported to be very efficient at producing genetically modified trees. Nonetheless, there are unresolved problems with plasmid sequences that remain in the plant genome. In this study, we demonstrated a DNA-free genome editing technique in which purified CRISPR/Cas9 ribonucleoproteins (RNPs) are delivered directly to the protoplasts of a hybrid poplar (Populus alba × Populus glandulosa). We designed three single-guide RNAs (sgRNAs) to target the stress-associated protein 1 gene (PagSAP1) in the hybrid poplar. Deep sequencing results showed that pre-assembled RNPs had a more efficient target mutagenesis insertion and deletion (indel) frequency than did non-assembled RNPs. Moreover, the RNP of sgRNA3 had a significantly higher editing efficacy than those of sgRNA1 and sgRNA2. Our results suggest that the CRISPR/Cas9 ribonucleoprotein-mediated transfection approach is useful for the production of transgene-free genome-edited tree plants.

• Korean Journal of Breeding Science
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• v.50 no.4
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• pp.424-433
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• 2018

Watermelon (Citrullus lanatus) is an economically important vegetable crop all over the world, which has functional compounds such as lycopene and citrulline. Gummy stem blight caused by Didymella bryoniae is one of the most devastative diseases in watermelon. Single nucleotide polymorphisms (SNPs), which are genetic variations occurring between individuals with respect to a single base, were often used to construct genetic linkage maps and develop molecular markers linked to a variety of horticultural traits and resistance to several diseases. In this study, we developed high-resolution melting (HRM) markers based on SNPs generated from NGS resequencing of two parents in watermelon. Plant materials were C. lanatus '920533' (female and susceptible parent), C. amarus 'PI 189225' (male and resistant parent), and their $F_1$ and $F_2$ progenies. A total of 13.6 Gbp ('920533') and 13.1 Gbp ('PI 189225') of genomic sequences were obtained using NGS analysis. A total of 6.09 million SNPs between '920533' and 'PI 189225' were detected, and 354,860 SNPs were identified as potential HRM primer sets. From these, a total of 330 primer sets for HRM analysis were designed. As a result, a total of 61 HRM markers that have polymorphic melting curves were developed. These HRM markers can be used for the construction of SNP-based linkage maps and for the analysis of quantitative trait loci (QTLs) related to gummy stem blight resistance.

• Journal of Korean Home Economics Education Association
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• v.16 no.3
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• pp.131-146
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• 2004

The Purpose of this study is to suggest strategies for environmental education through the Practical Arts(TechnologyㆍHome economics) Subject in a viewpoint of the clothing & textiles resources to resolve problems in the clothing life area. For this, this study was carried out through review of literature which is related with the consumption, the environmental problems, the environmental policies, and regulations of the government and new environmental technologies, of clothing & textiles industries and environmental education. The major findings of the study were as follows; 1) The environmental education system model in a viewpoint of the Clothing & Textiles resources was developed. This model system is consisted with interactions on school, government, industry, home and non-government organizations. Thus, the fact that Practical Arts(TechnologyㆍHome economics) Subject were the most effective subject to teaching the environmental education viewpoint of the Clothing & Textiles resources was confirmed. 2) The standards were analysed out to analyse the contents in the clothing area of the Practical Arts(TechnologyㆍHome economics) Subject. It were consist of 4 factors and 12 elements under the factors: Awareness of clothing & textile resources(clothing consumption, production of clothing & textile and environmental problems). Planning and buying of clothing(planning, buying), Management of clothing(understand of textile. human body & environment, laundering and Environmental pollution, arrangement & conservation) Recycling & exhaust of clothing(contribution, redesign, recycling, exhaust) 3) Analysing the current Practical Arts (TechnologyㆍHome economics) subject from the Environmental education in the clothing section, the environmental education related with clothing were taught the most in the middle school course, and environmental contents were concentrated in the recycling factors. but not so much on other factors. 4) After analysing the Practical Arts (TechnologyㆍHome economics) subject, the strategies were suggested for reinforcing the environmental education in the clothing of the Practical Arts(TechnologyㆍHome economics) subject.