水稻台稉9號癒傷組織及細胞培養之研發及其應用

Development and Application of Callus and Cell Culture of Oryza sativa var. japonica Taiken 9

蔡顓印(Chuan-Yin Tsai)

指導教授 ： 鄭秋雄

國立屏東科技大學/農學院/熱帶農業暨國際合作系所/碩士(2007年)

https://doi.org/10.6346/NPUST.2007.00013

摘要

本研究之目的首為研發水稻癒傷組織誘導培養、細胞培養及植株再生之培養系統，進一步應用於進行抗白葉枯病細胞系之篩選。以水稻台稉9號成熟種子為培植體，探討培養基、水晶洋菜含量、蔗糖含量及植物生長調節劑對誘導水稻台稉9號種子癒傷組織之影響。結果以含2 mg/l 2,4-D、3%蔗糖、0.4%水晶洋菜之CS-1培養基置於500 lux光照下培養45天之癒傷組織形成量最佳，單位培植體種子之癒傷組織鮮重約0.334 g。另，觀察2,4-D含量（0.5-5.0 mg/l）對癒傷組織顆粒大小與質地之影響，則顆粒大小會隨2,4-D含量之提高而逐漸變小，但球形顆粒狀、球形團粒與整體狀等質地在所有處理中均有發生。細胞培養則以含2 mg/l之2,4-D之CS-1培養基誘導45天之癒傷組織為材料，並懸浮細胞建立培養第10天之懸浮細胞數最多，每3 g鮮重癒傷組織可獲得約1.4×106個細胞。探討培植體癒傷組織鮮重、植物生長調節劑、光照度、水晶洋菜含量對癒傷組織植株再生之影響，結果以含2 mg/l之 NAA及4 mg/l之kinetin、3%蔗糖、0.6%水晶洋菜之CS-1培養基，並置於3500 lux光照下之植株再生率最佳，約60%。以上述培養細胞誘導條件所誘導之懸浮細胞為供試材料，以白葉枯病菌為篩選劑，利用懸浮細胞與白葉枯病菌共同培養法進行抗白葉枯病細胞系之篩選，將篩選之抗白葉枯病細胞系與水稻台稉9號培養細胞分別處理白葉枯病菌培養過濾液，結果其細胞殘存率分別為96%與50.4%，顯示篩選之抗白葉枯病細胞系對病菌培養過濾液具有抗性。抗白葉枯病細胞系連續繼代培養於含2 mg/l 2,4-D之CS-1培養基中可分化出細胞團，細胞團亦可分化出體胚，但體
胚無法持續成長為植株。

關鍵字

水稻台稉9號 ； 組織培養 ； 癒傷組織 ； 細胞培養 ； 植株再生 ； 白葉枯病菌 ； 白葉枯病 ； 抗病篩選 ； 抗白葉枯病細胞系

並列摘要

The purposes of this study are first to develop the culture systems of callus induction, cell culture and plantlet regeneration of rice, and further apply to the selection of bacterial leaf blight-resistant cell lines. Using the mature seeds of Oryza sativa var. Japonica Taken 9 as the explants, the effects of the media, contents of gelrite agar and sucrose and plant growth regulators on callus induction of seeds of O. sativa var. Japonica Taken 9 were evaluated. The results indicated that CS-1 medium containing 2 mg/l 2,4-D, 3% sucrose, 4% gelrite agar at 500 lux light intensity were the best in callus formation, the fresh weight of callus per seed explants about 0.334 g. To observe the effects of 2,4-D contents (0.5-5.0 mg/l) on granular size and texture of callus, granular size would gradually decreased with the increase of 2,4-D content, but callus texture of granular, granulate cluster and massive has been occurred in all treatments. In cell culture, using the callus cultured on CS-1 medium containing 2 mg/l 2,4-D for 45 days as the materials and the suspension cells establish cultured for 10 days could obtained the maximum suspension cells, which about 1.4×106 cells per 3 g fresh weight callus. To evaluate the effects of the fresh weight of callus per explant, plant growth regulator, light intensity and gelrite agar on plantlet regeneration of callus, CS-1 medium containing 2
mg/l NAA, 4 mg/l Kinetin, 3% sucrose and 6% gelrite agar at 3500 lux light intensity had the best result in the rate of plantlet regeneration, which about 60%. Using the suspension cells, induced from the above-mentioned conditions of cell culture, as the materials, Xanthomonas oryzae pv. oryzae as the selection agent and the method of suspension cells co-cultured with X. oryzae pv. oryzae to proceed the selection of bacterial leaf blight-resistant cell lines. The selected bacterial leaf blight-resistant cell lines and culture cells of Oryza sativa var. Japonica Taken 9 were treated with the culture filtrate of X. oryzae pv. oryzae, the survival rate of cells were 96% and 50.4%, respectively. The result indicated that the selected bacterial leaf blight-resistant cell lines were resistant to the culture filtrate of X. oryzae pv. oryzae. The bacterial leaf blight-resistant cell lines subcultured on CS-1 medium containing 2 mg/l 2,4-D could differentiate to clusters, and clusters could differentiate to somatic embryos, but couldn’t continue grew to plantlets.

並列關鍵字

Oryza sativa var. japonica Taken 9 ； tissue culture ； callus ； cell culture ； plantlet regeneration ； bacterial leaf blight ； Xanthomonas oryzae pv. oryzae ； disease-resistant selection ； bacterial leaf blight-resistant cell lines

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