초록 ▼

Ⅳ. 연구개발결과
(필요에 따라 제목을 달리할 수 있음)
1. 자체 개발한 벡터 시스템을 가지고 유전자발현 백터 제작
2. 충해저항성/제초제저항성/가뭄저항성/건강기능성 형질전환체 생산
3. 생산된 형질전환체 유전자 도입 확인 및 기능 분석 수행
4. 비타민a강화 콩 농업적 특성평가 및 고정계통 선발 - 농업적 특성이 안정적이고 생산성이 높은 계통 선발, 도입유전자의 고정계통 선발
5. 해충저항성 콩 농업적 특성평가 - 계통간의 농업적 특성에 대한 변이의 폭이 크고 계통 내에서도 변이가 큼으로 지속적으

Ⅳ. 연구개발결과
(필요에 따라 제목을 달리할 수 있음)
1. 자체 개발한 벡터 시스템을 가지고 유전자발현 백터 제작
2. 충해저항성/제초제저항성/가뭄저항성/건강기능성 형질전환체 생산
3. 생산된 형질전환체 유전자 도입 확인 및 기능 분석 수행
4. 비타민a강화 콩 농업적 특성평가 및 고정계통 선발 - 농업적 특성이 안정적이고 생산성이 높은 계통 선발, 도입유전자의 고정계통 선발
5. 해충저항성 콩 농업적 특성평가 - 계통간의 농업적 특성에 대한 변이의 폭이 크고 계통 내에서도 변이가 큼으로 지속적으로 안정화한 다음 고정계통 선발 필요
6. 애기장대 TP promoter를 이용하여 양방향 프로모터 제조
7. 제조된 양방향 프로모터를 이용하여 두 개의 유전자를 동시에 발현시킬 수 있는 벡터 시스템 제조 (pCKLSL-B)
8. pCKLSL-B 벡터에 병 저항성 유전자 (PepEST) 및 스트레스 저항성 유전자 (ABF3)를 발현시킬 수 있는 유전자 벡터를 제조하여 잔디로 도입시켜 형질전환체 확보
9. 잔디 형질전환체 분석을 통하여 잔디 식물체에서 양방향 프로모터가 작용함을 검증하였고, 도입된 유전자들에 의한 제초제 저항성을 포함하여 스트레스 저항성 및 병 저항성 표현을 검증
10. 따라서 양방향 프로모터를 이용한 pCKLSL-B 벡터 시스템을 이용하여 복합형질 식물체 개발이 가능함을 제시

Abstract ▼

Soybean is one of the most important crop in Korea due to its high nutritious value and frequent food consumption by most of Korean. Recently, domestic production of soybean can not meet the massive demand of soybean for food and industrial use for oil, protein, and various processed food. So far, c

Soybean is one of the most important crop in Korea due to its high nutritious value and frequent food consumption by most of Korean. Recently, domestic production of soybean can not meet the massive demand of soybean for food and industrial use for oil, protein, and various processed food. So far, conventional breeding took part of major role to breed new variety. However, biotechnology including genetic transformation is getting more important to satisfy many new demands from agricultural environment and industry.
In this study, we brought 3 BT toxin genes for soybean transformation to develop insect resistance soybean. Soybean transformation has been tried to produce enough transgenic plants with previously developed high-efficiency soybean transformation method. Our method was based on general soybean transformation method with cotyledonary node but minor modification was added to increase efficiency of genetic transformation for Korean soybean variety. We applied dipping/sonication/vacuum during soybean transformation and at least two of three additional treatments were used for soybean transformation. After transformation all material was advanced through shoot induction, shoot elongation, root induction, transfer to small port and large port in green house. PPT painting was primary selection for genuine transformant and DNA analysis was carried for confirmation of transgene, later on. To obtain copy number of transgene real-time PCR was used for T0 plant material.
In this study, two individual research teams have carried two separate experiments. In an experiment, all the transgenic soybeans produced in this project has cultivated in GMO field located at Gunwi province, Gyunbuk. All the T1 or T2 seeds were germinated green house first and transferred to field after PPT leaf painting to confirm gene introduction. Plotting density was 70 cm X 15cm, and 10 - 15 plants were transferred for each trangenic line. Investigation was performed on culm length, number of branch, number of pod, seed weight of 100, maturing period, and total weight of seed per plant.
Every year, field deployment was performed from middle of May to early June. Deployed soybean was harvested from middle of October to early November. Yield parameters were investgated during the harvest time or after harvest.
In an another study, physiological and molecular analysis including DNA analysis, gene expression, confirmation of gene copy number were determined. For this project, 3 BT toxin genes, Cry1Ac, Cry1-Ac M8, Cry1-Ac M15, mainly transformed into soybean, After real time PCR, Southern blot and flanking sequence analysis, about 10 lines of transgenic soybean plants, 4 Cry1Ac, 3 for Cry1-Ac M8, 3 for Cry1-Ac M15, respectively, putatively satisfied all the requirements and demands for single insertion event. These 10 lines of soybean transgenic plants are further analysing to obtain whole inserted DNA sequences. This year we will continue to survey their field performance in many agronomic traits.
In this study, the preliminary growth and test of Korean soybean cultivar Kwangan was primarily tried to compare those transgenic plants after gene transformation. Cultivation of Kwangan was carefully noted for their basic agricultural traits in the field. With previously produced soybean transgenic plant, PAC lines where the content of beta-carotene was greatly increased, selection has been conducted to find fixed homozygote lines among progenies. For those putative homogeneous fixed lines, DNA was analysed and seek for the flanking sequences. After 2-3 generation of field deployment, some segregating line with single copy of PAC gene was obtained. Through the further selection, finally some of the segregating transgenic lines were determined as a fixed and flanking DNA beside of T-DNA insertion was extracted from the lines. One line, 7-3-2-2-1, showed their flanking sequences exactly matched to soybean chromosome sequences (chromosome number 14, 30786587~30786732bp). As a result, we concluded that T-DNA was integrated into soybean chromosome #14 and intergenic position. There was no known gene within 20 Kbp according to blast search on DNA sequences.

목차 Contents

• 완결과제 최종보고서 ... 1
• 제출문 ... 2
• 요약문 ... 4
• SUMMARY ... 8
• 목 차 ... 10
• 제 1 장 서 론 ... 11
• 제 2 장 국내외 기술개발 현황 ... 12
• 제 3 장 연구개발수행 내용 및 결과 ... 15
• 제 1 절. 콩형질전환체 대량 생산(기본백터제작/클린식물체확인) ... 15
• 제 2 절 : 형질전환체 재배/ 증식 및 목표형질 생리분석 ... 79
• 제 3 절 : 형질전환체 포장검정 및 계통생산 ... 95
• 제 4 절 : 형질전환용 백터 시스템 개발 ... 109
• 제 4 장 연구개발목표 달성도 및 대외기여도 ... 128
• 1절 : 목표대비 달성도 ... 128
• 2절 : 정량적 성과(논문게재, 특허출원, 기타)를 기술 ... 132
• 제 5 장 연구개발결과의 활용계획 ... 141
• 제 6 장 연구개발과정에서 수집한 해외과학기술정보 ... 143
• 제 7 장 기타 중요 변동사항 ... 144
• 제 8 장 국가과학기술종합정보시스템에 등록한 연구장비현황 ... 145
• 제 9 장 참고문헌 ... 146
• 끝페이지 ... 152