細胞來源與培養方法對台稉九號水稻懸浮細胞增殖之影響

Effects of Cell Source and Culture Method on the Proliferation of Suspension Cells in Oryza sativa var. japonica ‘Taiken 9’

溫敏琪(Min-Chi Wen)

指導教授 ： 鄭秋雄

國立屏東科技大學/農學院/植物醫學系所/碩士(2014年)

https://doi.org/10.6346/NPUST.2014.00203

摘要

本研究目的為建立穩定之水稻懸浮細胞培養技術。為獲得增殖能力及分化能力強之懸浮細胞，以臺稉9號水稻成熟種子與其無菌苗之根為供試材料。比較種子與根來源癒傷組織之懸浮細胞數與細胞增殖率，結果以根來源較佳，其克鮮重癒傷組織之懸浮細胞數為2.4#westeur024#107cells/ml，較種子來源懸浮細胞者多出2.2倍。水稻癒傷組織依質地可分為硬質癒傷組織（種子來源）、鬆軟癒傷組織（根來源）及泥狀癒傷組織（根來源）。比較種子與根來源懸浮細胞之初代培養第8天，其細胞增殖率分別為-6%與34%，顯示根來源細胞優於種子來源細胞。重複建立培養可提高種子與根來源懸浮細胞之初代培養之增殖率，並可減緩繼代培養時，懸浮細胞數下降之現象。最後比較硬質癒傷組織、鬆軟癒傷組織及泥狀癒傷組織之懸浮細胞之細胞增殖率，結果以泥狀癒傷組織最佳，其初代與第一次繼代之細胞增殖率分別90.6%與15.2%。這些懸浮細胞持續培養至第二次至第四次繼代培養時，可分化出大量之minicalli，於50 ml液體培養基中分別有1700顆、2600顆與1065顆，顯示泥狀癒傷組織來源之細胞具有較佳之分化能力。綜合上述可知，培養細胞來源（器官培植體與癒傷組織質地）、培養細胞密度及建立培養次數對懸浮細胞增殖能力之影響至鉅。本研究最佳之培養條件為：(1)以發芽後10日齡之水稻無菌苗之根（含根尖組織）為培植體（1 cm），(2)培養於含2 mg/l 2,4-D之CS-1培養基，並置於無光環境下培養14天誘導之泥狀癒傷組織為細胞來源，可獲得增殖能力及分化能力優之水稻懸浮細胞。

關鍵字

水稻 ； 細胞增殖 ； 細胞來源 ； 泥狀癒傷組織 ； 建立培養

並列摘要

The purpose of this study was to establish the stable culture technology of suspension cells in rice. In order to get the suspension cells that have good abilities in proliferation and differentiation, the mature seeds and the root of in vitro seedling in Oryza sativa var. japonica cv. Taiken 9 were used as the materials. In comparison of the number and proliferation rate of suspension cells of callus induced from the seed and the root. Callus induced from the root had the best result in number of suspension cells per gram callus, about 2.4#westeur024#107cells/ml, which was 2.2-fold higher than the callus induced from the seed. Callus were grouped based on the texture, harden nodule callus (induced from seed), friable callus (induced from root) and sticky callus (induced from root). Compared proliferation rate of the suspension cells, induced from the seed and the root, after primary culture 8 days, which were 34% and -6% respectively, the result indicated that the culture cells induced from the root better than from the seed. Repeated establishment culture could raise proliferation rate of the suspension cell induced from the seed and the root in primary culture, in addition, it could slow down the decrease in the number of suspension cells in the period of subculture. In comparison of proliferation rate of the suspension cells among harden nodule callus (induced from seed), friable callus and sticky callus (induced from root). Sticky callus was the best, the primary culture and the first subculture were 90.6% and 15.2% respectively. When these suspension cells continued to culture the second to the fourth subculture, which could differentiated a vast amount of minicalli, there were 1700, 2600 and 1065 minicalli per 50 ml respectively. It’s found that the suspension cells induced from the sticky callus had the good ability in differentiation. Summarize the above-mentioned, these results indicated that source of the culture cell (organ explant and callus texture), cell density and times of establishment culture had a great influences on the abilities of proliferation and differentiation of the suspension cells. In this study, it’s found that (1) the root section (involve the root tip) of in vitro seedling, 10-day-old after germination, offered as the explants (1 cm), and (2) the induced sticky callus that cultured on CS-1 medium containing 2 mg/l 2,4-D at dark for 14 days as the source of culture cells were the best culture conditions, could obtain the suspension cells that have the good abilities in proliferation and differentiation in Oryza sativa var. japonica cv. Taiken 9.

並列關鍵字

rice (Oryza sativa var. japonica cv. Taiken 9) ； cell proliferation ； cell source ； sticky callus ； establishment culture

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