초록

본 연구에서는 Agrobacterium rhizogenes를 이용하여 자초(Lithospermum erythrorhizon) 세포를 형질전환 시켜 그로부터 모상근(hairy root)를 유도하고, 유도된 모상근의 배양 및 shikonin 생산을 위한 새로운 형태의 실험실 규모의 생물반응기를 설계하고 그 응용성을 검토 하는 것을 주내용으로 하였다.

Abstract ▼

The use of a bioreacter system for the production of plant secondary metabolite was examined in this study. Lithospermum erythrorhizon cells were transformed by using A. rhizogenes, and their growth and shikonin productivity were investigated.Hairy roots, the transformants of L. erythrorhizon, were

The use of a bioreacter system for the production of plant secondary metabolite was examined in this study. Lithospermum erythrorhizon cells were transformed by using A. rhizogenes, and their growth and shikonin productivity were investigated.Hairy roots, the transformants of L. erythrorhizon, were effectively induced by A. rhizogenes. The transformation was confirmed by the opine analysis. Of twenty hairy root clones, a clone designed as HR 9, which showed the highest growth was selected and used in the present study. In a preliminary flask culture, it was observed that Schenk and Hildebrandt medium diluted triple times(one-third salt concentration) was most effective for both root growth and shikonin production;it was chosen as the basal medium.By using the basal medium, effects of the concentration of sucrose, potassium nitrate, ammonium phosphate and cupric sulfate on shikonin production were examined. Their optimal concentrations were found to be;40g/l sucrose, 2.5g/l potassium nitrate, 100mg/l ammonium phosphate and 0.25mg/l cupric sulfate.Using the modified Schenk and Hildebrandt medium, shikonin was produced at a concentration of 320mg/l in the flask culture for 24 days. In the bubble column bioreactor and in situ extractive bioreactor product titre attained were 109mg/l/24 day and 258mg/l/35 day, respectively. Although the product titre was comparatively low, it should be emphasized that the use of bubble column bioreactor and in situ extractive bioreactor for mass culture of hairy root could offer a possible advantage; extended production phase. More detailed study including futher improvement of cell line and optimal operation of the bioreactor system is considered necessary to enlighten the practical application of the bioreactor system of hairy root culture.

목차 Contents

• 제1장 서론...11
• 제2장 실험재료 및 방법...13
• 제1절 식물재료 및 균주...13
• 제2절 모상근 유도...13
• 제3절 배지 및 배양방법...14
• 제4절 유동충 생물 반응기...14
• 제5절 분석방법...14
• 1. 모상근 증식...14
• 2. 당 정량...14
• 3. Shikonin 정량...18
• 4. Opine 분석...18
• 5. 기타...18
• 제3장 결과 및 고찰...20
• 제1절 모상근 유도...20
• 제2절 모상근 세포주 선발 및 형질전환 유무 확인...20
• 제3절 모상근 생육 및 shikonin 생산을 위한 배지 선발...20
• 제4절 Shikonin 생산을 위한 배지조성 검토...25
• 1. 모상근 증식 및 shikonin 생산에 미치는 탄소원의 영향...25
• 2. 모상근 증식 및 shikonin 생산에 미치는 무기염류의 영향...25
• 제5절 모상근 배양에 의한 shikonin 생산...30
• 1. 프라스크에서 모상근 배양...30
• 2. Bubble column reactor에서 모상근 배양...30
• 3. 동시추출 생물반응기 디자인 및 shikonin 생산...37
• 제4장 요약...40
• 참고문헌...41