人蔘의 組織培養에 있어서의 培地成分 및 形態發生에 관한 硏究

李載斗,李相泰 成均館大學校 1970 論文集 Vol.15 No.-

Tissues of Panax Schinseng Nees root were cultured on the synthetic agar media to investigate the nutrient efficiency on the callus induction and the organ formation And the differentiation pattern of the callus mass and the structure of the induced organ(root) were observed internally. On White's medium, callus formation needed the supplement of 2,4-D(5mg/l) and kinetin(1.0mg/l), and on Ms medium the root induction NAA(0.2mg/l) and kinetin(0.1mg/l). In order to investigate the effect of inorganic components on callus formation, the inorganic part of White's medium was substituted with those of Heller, Murashinge and Skoog, and Earle. As the result of culture Earle's was most effective. On the other hand, the roots were induced from the meristem in the deep region of callus mass. And this meristem is similar to the pericambium of tap root, so they are same on the pattern of morphogenesis.

細胞培養에 따른 人蔘 核型異常

李載斗,朴鍾範 成均館大學校 1982 論文集 Vol.31 No.-

This study was the results to examined the variation of chromosome number in the suspension culture of Korea ginseng (Panax ginseng) callus cell was added various growth substance. Basic medium was Murashige & Skoog's solution and growth substance used in this experiment was 2,4-D, kinetin and 2,4 D+Kinetin. Concentration of the growth substance was 10^-4 M, 10^-6 M and 10^-8 M respectively. Suspension culture was carried out in gyratory shaker (16rpm) and kept with 25+1℃. The effects of concentrations of growth substance was in the frequency of cell division was higher in medium added to 2, 4 D in 10^-4 M concentration, and the degree of aberration of chromosome number was higher in above medium also. In medium added to kinetin the both of frequency of cell division and the degree of the aberration of chromosome number was lower, otherwise, in medium added 2, 4 D-kinetin in 10^-6 M concentration the frequency of cell division and the degree of the aberration of chromosome number were very higher than other media culture. Also, when we observed the chromosome aberration in above various media in different concentration normal diploid kayrotype (2n=48) were observed in media added to 2, 4 D-kinetin in 10 ^-4 M and 10^-6 M concentration in large number, but the other media the tentency toward aneuploidy were observed. In this experiment, it is suggested that growth substance, espicially 2, 4-D, in suspension culture may be responsible for consderable chromosome aberration.

인삼 원형질체의 세포벽 형성에서 Fibril형성에 관한 전자현미경적 연구

이재두,김동균 성균관대학교 기초과학연구소 1990 論文集 Vol.41 No.1

인삼 callus 원형질체를 실험재료로 사용하여, 세포벽 재형성과정 중에서 fibril형성을 광학 및 전자현미경으로 조사하였다. 세포벽 형성 물질이 들어있는 vesicle들은 활성화된 dictyosome에서 많이 만들어 지며 exocytosis에 분비되어, 원형질체 표면에서 fibril이 형성된다. The formation of cell wall fibrils was studied by light microscope and electron microscope on the surface of isolated callus protoplasts from Korean ginseng(Panax ginseng C. A. Meyer). Vesicles which originated from active dictyosome were migrated out of cell by exocytosis and then fused to form fibrils. We observed that the fibrils had not been formed on incubated protoplasts until a lag period was elopsed. After 5-6 hours, many of short fibrils were formed 20-30 nm in width. These fibrils were elongated and randomly interlaced.

植物組織培養에 있어서 生長物質의 效果에 關한 硏究

李載斗,洪性式 成均館大學校 1978 論文集 Vol.25 No.-

The object of this study is to clarify the effects of single and combination treatments of α-naphthalene acetic acid (NAA), 2,4-dichlorophenoxy acetic acid (2,4-D), indole-3-acetic acid (IAA) and kinetin in young stem cultures of Paeonia albiflora. Basic medium was Murashige & Skoog's medium, and all media were adjusted to pH 5.8 before autoclaving. This experiments were carried out in the dark room and it was kept with temperature of 25±1℃. In single treatment of above various growth substances, the optimum concentrations on callus initiation were NAA of 0.5mg/l,2,4-D of 0.5mg/l,IAA of 0.05mg/l and kinetin of 0.3mg/l. In combination treatments of above various growth substances 2,4-D+kinetin would be very efficient to clone formation, but did notinduce root differentiation. And, it's suggested that the effects of 2.4-D+kinetin+NAA, 2,4-D+kinetin+IAA and 2,4-D+kinetin+NAA+IAA were same as 2,4-D+kinetin, and NAA+2,4-D showed clone formation efficiently. Also, adding of NAA or 2,4-D established clone formation and root differentiation. On the other hand, NAA+kinetin would be effective to root differentiation. Nevertheless, excellent root differentiation and development would be established from the clones in which was trasfered from basic medium+NAA (0.5mg/l) to basic medium.

細胞培養에 依한 組織發生에 관한 硏究 : I. 植物生長物質이 分裂組織과 髓細胞의 組織發生에 끼치는 影響에 對하여 I. Effect of plant growth substance for histogenesis of meristematic cell and pith cell

李載斗,金昌基 成均館大學校 1968 論文集 Vol.13 No.-

은행나무(Ginkgo biloba)의 分製組織과 髓에서 切取한 細胞群(cell colonies)을 indol-3-acetic acid (IAA, 0.1 ppm), gibberellic acid(G.A. 0.025 ppm), IAA+GA(1:1) 및 glutamic acid (300 ppm)을 15% coconut milk 및 2% sucrose 添加 White 氏 培養液에 各各 添加시킨 培養區에서 7日, 14日, 21日間 培養한 바 다음과 같은 結果가 나타났다. 첫째로 모든 培養區중에서 IAA 添加區가 가장 活潑한 組織發生을 하고 있으며 그 다음이 GA區로서 이 區에서는 髓細胞의 發生이 더욱 顯著함을 관찰할 수가 있으며 IAA+GA添加區는 單獨處理區에 比하여 低調한 發生相을 보이고 있어 組織培養시와 마찬가지로 相互制御作用이 있음을 엿볼 수가 있다. 室素源으로서의 glutamic acid 添加區는 若干의 發生相을 보이나 植物生長物質의 添加區에 比하여 顯著하게 低調한 發生相을 보여 주고 있다. We have been studied on the growth and the developement rate of the cell colonies obtained from the meristem and pith of Ginkgo biloba by cultivating in vitro during 7, 14 and 21 days. The culture media were White's solution adding 15% coconut milk and 2% sucrose, and each medium was mixed with Indol-3-acetic acid(IAA), Gibberellic acid(GA), IAA+GA and Glutamic acid, respectively. It has been found that the growth and the development of the cell colonies was most outstanding in the medium adding IAA, and in the medium adding GA the developement of pith cell was outstanding especially. However, the developement phase in IAA+GA was less outstanding as compared with solitary treatment of IAA and GA, therefore, it seemed that there would exist mutual inhibition phenomena between IAA and GA. In the medium adding glutamic acid as nitrogen source, showed some developement, but the rate of the growth and the developement was less than that in the medium adding plant growth substances.

人蔘(Panax ginseng C.A. Meyer)의 胚培養에 관한 硏究 : Ⅰ. Protocorm-like body의 形成 Ⅰ. Formation of Protocorm-like body

李載斗,朴鍾範 성균관대학교 기초과학연구소 1985 論文集 Vol.36 No.1

The growth and differentiation of the embryo isolated from the Korean ginseng. Panax ginseng C.A. Meyer, cultured in the medium in vitro have been investigated by the light microscopic examination of the developmental characteristics of organprimodium and the patterns of rooting and shooting. The embryos, excised from the post-maturated seeds, were cultured on the Murashige & Skoog(MS) medium containing various amount of 2,4-dichlorophenoxy acetic acid (2, 4-d) and kinetin under the 12L : 12D) photoperiod, and completely dark condition. The protocorm-like body developed only when embryos were taken from the seedling two weeks before the germination, and cultured in the dark in the presence of 10^-6M 2,4-D and 10^-8M kinetin. Under such conditions a callus mass developed after 15 weeks and a protocorm-like body developed after 5 months, which eventually developed to become root. Microscopic observation of the sections of pre-protocorm-like body revealed a meristematization in both parenchyma cells in the center of cortex and the vascular cambium cells, which subsequently developed into nodules prior to the development or protocorm-like body.

高麗人蔘에 育種의 關한 硏究 : Panax ginseng C. A. Meyer

李載斗,洪性式 成均館大學校 1980 論文集 Vol.27 No.-

This study is the report of karyotype analysis in somatic cell and the investigation of chromosomal aberration was induced by growth substances, X-ray and colchicine as mutagen in tissue culture of Korean ginseng. We obtained the following results. For the karyotype analysis, we used embryo in the seedling and shoot bud, the number of chromsomes(2n) is 48. Of these 46 make pair showning that these are A-chromosome, the homologous chromosome are identified from the centromere and chromosome band and rest of them which correspond to B-chromosome did not form pair corresponds to B-chromosome. It is interresting that one of the B-chromosome is the largest and the other is the smallest. For the induction of artificial mutation, we used growth substances as 2,4-dichlorophenoxy acetic acid(2,4-D) and cytokinin, and X-ray and colchicine. This treatment induce the variety of chromosome number. Sample cultured in Murashige-Skoog media contained growth substances show mostly diploid chromosome and there were some haploid (n=24) and aneuploid. Among the aneuploid observed are 2n-2, 2n+4 or 2n+12. In addition to this, there were tetraploid (4n=96) and their aneuploid (4n-4, 4n-1 and 4n+6). However, X-ray irradiation cause n and 4n disappear and some aneuploid of 2n type (2n-8, 2n-2 and 2n+2) and 2n phase. Next, we observed the effect of colchicine in the suspension culture, in this experiment we recognized n, 2n and their aneuploid. However, we did not observed tetraploid. The variation from n to 2n is gradual and there is no 4n indicating that the furture experiment of cytological mechanism of mutation induced by growt-h substances will be useful of developing new breed of Korean ginseng.

은행나무의 組織培養에 따른 器官形成에 關한 硏究

李載斗 成均館大學校 1976 論文集 Vol.23 No.-

This investigation was studied for various factor effected to the clone formation and the organogenesis of Ginkgo biloba in vitro. The media used this experiment was improved White's solution (1943) supplimented with 2, 4-dichlorophenoxy acetic acid (2, 4-D), α-naphthalene acetic acid(NAA), kinetin, adenine sulfate and casein hydrolysate, and was observed for effects on this supplementive substance in cultures respectively. Also, in order to pursue influence of light illumination in culture, made dark, dark versus light and light treatments in culture. Consequently, optimum the clone formation and the organogenesis from the piece of the stem apex and the cotyledonary node portion of Ginkgo biloba used as the most suitable materials was obtained on improved White's solution supplemented with α-naphthalene acetic acid (2mg/1), kinetin (0.025%), adenine sulfate(3mg/1) and casein hydrolysate (600mg/1). And under dark versus light clone and organ formation was much faster as compared to the dark culture. Under above culture condition the callus induction was after cultured during 7 days, and the rooting from the clone was obtained after cultured during 30 days, the shooting made after cultured during 40 days, therefore, was made plantlet.

炭素源과 은행나무 切斷胚軸의 生長 및 分化에 關한 硏究

李載斗 成均館大學校 1966 論文集 Vol.11 No.-

은행나무(Ginkgo biloba L.) 胚軸의 根端을 3 ㎜의 길이로 切斷하여 寒天添加無糖 Whit氏 培養液에 sucrose 2%(0.06M), 4%(0.117M), 8%(0.234M), 16%(0.468M), 및 glucose 2%(0.111M), 4%(0.222), 8%(0.444M), 16%(0.888M)을 넣고 各各을 試驗區로 하여 切斷根을 培養하였다. 各驗試區에는 材料 60個씩 使用하고 各各 7, 14, 21日間 培養한바 그結果는 다음과 같다. 切斷根端의 길이의 生長은 2% sucrose 21日區의 驗試區에서 가장 컸고 2%, 4%, 8%, 및 16%의 順序로 低下된 生長을 나타내었다. 그러나 切斷面에서 생기는 callus의 發生은 4% sucrose區에서 가장 旺盛하였으며 同時에 그의 組織分化의 進行도 또한 이 區에서 빨리 나타나고 있다. callus의 組織分化의 進行은 2%區와 4%區에서 生長樣相과 엇갈리는 現象을 보이고 있어 2%區에서는 組織分化의 進行이 늦은 callus의 形成이 比較的 旺盛하나 4%區에서는 組織分化의 進行이 빠른 callus의 形成이 旺盛하게 일어나고 있다. 그런데 8%區는 生長과 分化가 平行하게 나타나고 比較的 着實한 進行樣相을 보이나 16% glucose區에서는 生長과 分化가 아울러 大端히 낮은 樣相을 보이고 있다. 以上에서 알려진 事實로 根端分裂組織의 活動은 2%區에서 좋았고 callus의 形成 및 分化의 樣相은 4%區에서 優秀하였다. 特히 16% glucose區에서 生長 및 分化가 뚜렷이 低下된 原因은 이 區가 가장 높은 mole濃度이기 때문에 吸收가 잘 되지못하여 生長 및 分化에 必要한 炭素源의 不足으로 일어나는 現象이라고 보고 그의 培養에서 糖의 吸收量을 調査한 결과 다른 驗試區에 比하여 越等하게 낮았고 따라서 生體속의 消費量도 大端히 적었다. 다음에 培養液의 pH 變化는 培養日數에 따라서 酸性化하는 傾向을 띄고 있는 本實驗에서의 變化幅은 5.80∼3.75이다. 이 變化幅內에서는 糖類의 吸收의 支障이나 生長 및 分化에 對한 抑制作用이 일어나지 않음을 알수가 있었다. 結局 生長과 分化에 對한 糖類의 影響은 各各 糖類의 種類나 濃度에 따라서 差異가 나타나고 있으며 같은 mole 濃度에 있어서 sucrose는 glucose보다 더욱 좋은 炭素源이 됨이 確實하다. Excised roots (3㎜ section from root tip) of Ginkgo embryo was cultured in White's solution (agar added) supplemented sucrose 2, 4, 8, 16% and glucose 2, 4, 8, 16% respectively and each cultured during 7, 14, 21 days. In this experiment the results were summarized as follows. 1) The excised root has been enlarged to its maximum length in 2% sucrose medium during 21 days, and in sucrose medium growth rate decreased according to the order of 2, 4, 8, 16%, but in glucose medium growth rate decreased according to the order of 4, 2, 8, 16%. 2) The tissue differentiation of cell has appeared in contrast with growth rate and most active in 4% sucrose medium, but both of the differentiation of tissue and growth were seen less in 16% glucose medium. 3) The amount of sugar absorption from solution and consumption in materials were almost similar in sucrose and glucose medium, but in 16% sucrose medium, it was seen remarkably less, and the growth in length and titissue differentiation of tissue immediately. 4) The effect of sugars as a carbon source on growth and differentiation of tissue depended on concentration and kind of sugar. 5) The hydrogen ion concentration has been changed within range from pH 3.75 to 5.80 according to the days that it has been cultured. Acidities increased according to the length of the period that it has been cultured. Increase in acidity depended on how long it has been cultured.

根 形成에 對한 糖과 生長物質의 影響

李載斗,金昌基 成均館大學校 1966 論文集 Vol.11 No.-

은행나무(Ginkgo biloba L.)胚의 根端에서 3㎜를 切斷除去한 나머지의 shoot部分을 寒天添加無糖 White氏 培養液 (1943年)에 sucrose 2%, 4%와 IAA(0.001 ppm)와 IAA 및 GA(0.00025ppm)를 添加한 6種類의 培養液과 無糖液을 合한 7個의 試驗區에서 14日間 培養實驗하고 各試驗區에서 자란 根端切除胚의 切斷面에 가까운 pericycle 部位에서 側根이 發生하는 樣相을 調査하였다. 그 調査結果는 다음과 같다. (1) IAA 添加 4% sucrose區에서 根形成이 가장 旺盛하였으며 IAA+GA 添加 sucrose區에서 가장 低調하였다. (2) IAA 添加 2% sucrose區에서는 pericambium이 나타나나 아직 根形成이 뚜렷하지 않고, 2% sucrose區와 IAA+GA 添加 sucrose區에서는 全然 pericycle조차 나타나고 있지 않는다. (3) 糖添加培養에 IAA 添加를 함으로써 組織分化가 顯著하여지고 있으며 이것은 糖과 IAA가 根形成에 相乘作用을 하는 現象으로 볼수 있다. (4) GA+IAA 添加區는 IAA 添加區보다 生長 및 分化가 低調하였다. The shoot portion of embryo removed 3㎜. from root tip of Ginkgo biloba was cultured in the White solution containing agar, 2 and 4% sucrose was also added to this medium with IAA. (0.001 p.p.m.) and GA.(0.00025 p.p.m.). In this experiment, author investigated that the lateral root was developed from pericycle near the sliced root tip of embryo, which was grown in each medium during 14 days at 25±1℃. The results are as follows. 1. The 4% sucrose medium supplemented IAA. was most active one, supporting the root formation, but the other with IAA+GA. was inactive one. 2. Even though pericambium was developed in the 2% sucrose medium containing IAA., perieycle could not be found in the 2% sucrose medium containing IAA.+GA.. 3. When IAA. supplemented sucrose medium, tissue differentiation was marked. 4. The root formation and root differentiation was inactivated in sucrose medium containing IAA.+GA..