Abstract: The protoplasts of three Oryza species (O. rufipogon, O. glumaepatula and O. latifolia) were successfully cultured by using agarose-bead method in simplified protoplast culture medium (SPCM) with or without the nurse cell liquid through cell suspensions from mature embryos. Two of them (O. rufipogon, O. glumaepatula) regenerated green plantlets from protoplast-derived calli. The nurse cell liquid covering agarose-bead protoplasts could promote cell division and significantly increase the frequency of cell division (33.2%) and colony formation (19.0%). The hardening for protoplast-derived calli on the suitable proliferation media greatly benefited green plantlet regeneration. In the course of callus hardening, L3 medium showed the highest efficiency (32.1%) in the formation of embryogenic calli and embryo-like structure, thus resulting in producing moregreen plants.

Key words: nurse cell liquid , plant regeneration, proliferation medium, protoplast culture, wild Oryza species

摘要： 三个野生稻（O. rufipogon，O. glumaepatula 和O. latifolia）成熟胚，经愈伤组织诱导和悬浮细胞培养，分离原生质体，利用简化原生质体培养基（SPCM），结合琼脂糖包埋，并附加看护细胞培养液，这三个野生稻原生质体均得到了成功培养，其中两个（O. rufipogon和O. glumaepatula ）分化了绿色植株。看护细胞液在野生稻原生质体培养中，可促进细胞分裂，提早细胞分裂始期，细胞分裂频率（3.2％）和植板率（19.0%）分别比未加看护细胞液的培养方法高出13.3%和6.2%。看护细胞液可大大减少野生稻愈伤组织在液体中培养始期的死亡率，提高建立其悬浮细胞的成功率。原生质体克隆在适当的后培养基上进行培养，可以显著改良其结构，明显促进绿苗分化。尤其是L3培养基诱导原生质体克隆形成胚性愈伤组织或类似胚状体结构的频率达32.1%，显著高于MS和N6培养基，从而导致较高绿苗分化频率。

关键词: 野生稻, 原生质体培养, 植株分化, 看护细胞液