24个水稻抗稻瘟病单基因鉴别品系悬浮细胞系的建立

The establishment of cell suspension culture of 24 rice monogenic lines of IRRI-Japan

以IRRI-Japan24个水稻抗稻瘟病单基因鉴别品系和对照普感品种丽江新团黑谷（LTH）的成熟种胚为材料诱导愈伤组织，诱导过程中发现不同基因型愈伤组织诱导能力差异极大，愈伤组织诱导率在14．3％-97．1％，将2,4-D浓度从3．0mg／L提高到5．0mg／L时，可提高愈伤组织诱导率。经诱导成功的水稻愈伤组织继代培养一段时间后，不同基因型的愈伤组织继代特性出现明显差异，选取生长状态良好的单株系愈伤组织建立了水稻悬浮细胞系。悬浮培养过程中对胚性悬浮系的细胞形态变化进行了观察。发现细胞的形状由开始的不规则、长条形逐渐转交成近椭圆形、椭圆形；细胞质由很淡、透明、具明显的大液泡逐渐转变成浓厚、不透明、较小的液泡。大多数品系的单细胞率由30％-40％提高到60％～90％。本研究通过努力，建立了14种单基因型水稻材料的稳定悬浮细胞系，构建过程中发现基因型、继代培养特性、愈伤组织类型选择是影响胚性悬浮细胞系建立的重要因素。该悬浮细胞系的建立为进一步研究稻瘟菌分泌蛋白与水稻细胞互作的生化机理奠定基础。

Using mature embryos of 24 rice monogenic lines of IRRI-Japan and universally susceptible japonica rice variety Lijiangxintuan- heigu （LTH） as materials to induce callus, the capabilities of callus induction were rather various among different rice genotypes, of which the callus induction rate varied from 14. 3 % to 97.1%. The concentration of 2,4-D had significantly influence on callus induction rate, and higher callus induction rate could be achieved by increasing 2,4-D concentration from 3.0 to 5.0mg/L. When cultivated the callus on subculture medium for about 20 days or more,the well-grown calli from each same embryo were singled out to build the cell suspension lines of rice. The shape of the suspension cell was observed during the process of cultivation. At the forepart of the culture, the shape of the cells were anomal and strip, their vacuole and cytoplasm were big and rarefaction respectively. At the metaphase of culture, the shape of the cells became ellipse gradually, the vacuole and cytoplasm became small and dense. Finally,we established 14 genotypes of the cell suspension lines from 24 rice monogenic lines and one control variety in our experiments. During cell suspension culture establishment,the main technical factors of the establishment of cell suspension culture of rice line were discussed. The experiment indicated that genotype, the selection of callus and methods of subculture could greatly affect the qualities of cell suspension lines and also play an important role in if one monogenic line was suitable to liquid culture. The established 14 cell suspension lines could be used as essential materials in study on interaction mechanism of secreted proteins of Magnaporthe grisea and rice cells.