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담배(Nicotiana glauca) 카루스로 부터 Protoplast 및 Subprotoplast의 分離
呂邑東,蘇雄永 전북대학교 유전공학연구소 1988 遺傳工學硏究所報 Vol.1 No.1
As tools for the genetic modification of plants, the protoplasts were isolated from the green callus of Nicotiana glauca and were partitioned to subprotoplasts by gentle press operation. The isolation of protoplasts was taken for 5 hours in the enzyme solution(cellulase Onozuka R-10, 2%, macerozyme R-10, 1%, D-mannitol, 13% pH, 5.8)on the shaking bath(30℃, 70 strokes/min). The mean diameter was 45. 47±23.28μm and the mean volume was 4.92×10^-8 ㎤. The vacuoles was more easily observed by means of the deletion of cytoplasm and the destrution of cytoplasmic strands. After treated for 5 hours with the same enzyme solution for isolation of protoplasts, the callus clusters were pressed for 5 minutes by pincette through cover glass on them. The protoplasts were partitioned to about five subprotoplasts containing miniprotoplasts and cytoplasts. The ranges of diameter of miniprotoplasts and cytoplasts were 8.53-46.18μm and 8,58-52.18μm, respectively. One protoplast was partitioned to miniprotoplasts and cytoplasts at the ratio of 1:3 by the mean volume of protoplasts plsamolysed. But the observed ratio of miniprotoplast to cytoplast was 1:0.44. This result indicates that cytoplasts were more easily destructed than miniprotoplasts containing nucleus.
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담배 綠色과 白色 Callus로부터 分離된 原形質體의 融合 및 培養
呂邑東,蘇雄永 全北大學校 基礎科學硏究所 1983 基礎科學 Vol.6 No.1
To investigate the colony formation activities of the fused protoplasts, the protoplasts isolated from green and white calli of tobacco (Nicotiana glauca) were fused by the PEG treatments and cultured in MS medium. The satisfactory yields (90%) of isolated protoplasts in white and green calli were taken 4 and 5 hours, respectively. The rates were depended upon the 40% PEG (MW,4,000) volumes added to 100 μ1 protoplats (about 4×10^5/ml). The most effective volume, 100 μ1 PEG exhibited the high fusion rate (37%). The fused protoplasts suspended in MS medium supplemented with 2,4-D 3mg/1, kinetin 0.5mg/1 and 0.4M glucose as osmoticum were cultured. Their first and second cell divisions were observed on the 2nd and 4th day, respectively. And their colonies were observed on the 7th day.
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칼라코에 줄기切片의 器官分化에 미치는 生長調節物質의 影響
李康燮,呂邑東,蘇雄永 全北大學校 基礎科學硏究所 1991 基礎科學 Vol.14 No.1
In order to elucidate the effects of growth regulators on organogenesis in the stem explants of Kalanchoe daigremontiana, we have cultured the explants for 10 weeks on the MS medium with or without ammonium nitrate, supplemented with simultaneous combinations of auxin(IAA), gibberellin(GA_3) and cytokinin(BAP)The cocentrations of these growth regulators were 0.001-10.0 ㎎/l, respectively. The callus formation and organogenesis in the explants are mainly achieved under the simultaneous actions of the auxin and cytokinin. The optimal combination of growth regulators for callus formation was IAA 10.0 ㎎/l, GA_3 0.001 ㎎/l and BAP 0.1 ㎎/l. The optimal combinations for the adventitious root and bud formation are IAA 10.0 ㎎/l, GA_3 0.01 ㎎/l, BAP 0.01 ㎎/l and IAA 0.1 ㎎/l, GA_3 0.01 ㎎/l, BAP 1.0 ㎎/l, respectively. On the MS medium without ammonium nitrate, the callus and adventitious bud formation in the explants are better than those on the MS medium with ammonium nitrate, but the adventitious root formation is worse. The correlation between callus formation and organogenesis is negative on the both media.
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