石刁柏组织培养中体细胞胚发生的组织细胞学观察

INVESTIGATION ON HISTOLOGICAL CYTOLOGY OF THE SOMATIC EMBRYOGENESIS DURING TISSUE CULTURE OF ASEPARAGUS OFFICINALIS

以石刁柏(Asparagus officinalis)无菌苗的嫩茎切段为外植体,在含有1 mg/LNAA+0.5 mg/L BA的MS培养基上可100%地被诱导形成愈伤组织,在此条件下可长期继代,将继代的愈伤组织转入含有2 mg/L 2,4-D 0.5mg/L NAA的MS培养基上后,约有70%的愈伤组织块转变为胚性愈伤组织,这些胚性愈伤组织在3,4-D浓度进一步降低为0.5mg/L的条件下发育形成体细胞胚。切片观察表明:这些胚性愈伤组织是从愈伤组织的表层或近表层产生的。这些细胞核大,多核仁,细胞质浓、染色深的胚性细胞中的一些单个细胞处于与邻近细胞隔离状态,细胞壁也明显加厚。这些单细胞开始分裂,第一次分裂多为不均等分裂,形成一个大的基细胞和一个小的顶细胞,进一步分裂形成三细胞、四细胞、五细胞和具胚柄的多细胞原胚。原胚发育形成球形胚、梨形胚、香蕉形胚,由于在胚的一侧细胞分裂旺盛形成单子叶突起,最后形成子叶胚。其发育过程类似于单子叶植物合子胚的发育过程。

Callus originated from the tender stem of Aseparagus officinalis seedling which were placed on MS medium with lmg/L NAA and 0.5mg/L BA. The long subculture at this condition was permissible. When the subcultured callus werc transfered to MS medium with 2mg/L 2, 4-D and 0.5mg/L, embryogenic callus were induced from 70% of the callus, further decreasing the level of 2,4-D to 0.5 mg/L, somatic embryos were formed. Observing the paraffin section showed that the embryogenic callus were initiated from the epidermal or sub-epidermal layers of the callus. The single cell among the embryogenic cells which had bigger nuclei, multinucleoli, thicker cytoplasm and denser colour got isolated from each other, the wall became thicker, then they began to mitosis. The first time was unequal so that a bigger basal cell and a smaller apical cell were formed. More division were carried out, and the three-cell, four-cell, five-cell and multicellular proembryos were formed. The proembryo continued to develop into globular embryo, pear-shaped embryo and banana-shaped embryo. Due to the cells on one side of embryo divided rapidly, monocotyledon raised, and the cotyledonary embryo was formed. This kind of development was similar to that of the zygotic embryo in monocotyl.