He Yu-Chi, He Yu-Qing, Qu Liang-Huan, Sun Meng-Xiang, Yang Hong-Yuan
Key Laboratory of MOE for Plant Developmental Biology, College of Life Science, Wuhan University, Wuhan 430072, China.
Plant J. 2007 Feb;49(3):515-27. doi: 10.1111/j.1365-313X.2006.02970.x.
We have developed a reliable in vitro zygotic embryogenesis system in tobacco. A single zygote of a dicotyledonous plant was able to develop into a fertile plant via direct embryogenesis with the aid of a co-culture system in which fertilized ovules were employed as feeders. The results confirmed that a tobacco zygote could divide in vitro following the basic embryogenic pattern of the Solanad type. The zygote cell wall and directional expansion are two critical points in maintaining apical-basal polarity and determining the developmental fate of the zygote. Only those isolated zygotes with an almost intact original cell wall could continue limited directional expansion in vitro, and only these directionally expanded zygotes could divide into typical apical and basal cells and finally develop into a typical embryo with a suspensor. In contrast, isolated zygote protoplasts deprived of cell walls could enlarge but could not directionally elongate, as in vivo zygotes do before cell division, even when the cell wall was regenerated during in vitro culture. The zygote protoplasts could also undergo asymmetrical division to form one smaller and one larger daughter cell, which could develop into an embryonic callus or a globular embryo without a suspensor. Even cell walls that hung loosely around the protoplasts appeared to function, and were closely correlated with the orientation of the first zygotic division and the apical-basal axis, further indicating the essential role of the original zygotic cell wall in maintaining apical-basal polarity and cell-division orientation, as well as subsequent cell differentiation during early embryo development in vitro.
我们已经在烟草中建立了一个可靠的体外合子胚胎发生系统。借助一个共培养系统，双子叶植物的单个合子能够通过直接胚胎发生发育成可育植株，在该共培养系统中，受精胚珠被用作饲养层。结果证实，烟草合子能够按照茄科类型的基本胚胎发生模式在体外分裂。合子细胞壁和定向扩张是维持顶端 - 基部极性以及决定合子发育命运的两个关键点。只有那些具有几乎完整原始细胞壁的分离合子能够在体外继续有限的定向扩张，并且只有这些定向扩张的合子能够分裂成典型的顶端和基部细胞，并最终发育成带有胚柄的典型胚胎。相比之下，去除细胞壁的分离合子原生质体能够增大，但不能像体内合子在细胞分裂前那样定向伸长，即使在体外培养过程中细胞壁再生也是如此。合子原生质体也能进行不对称分裂，形成一个较小和一个较大的子细胞，它们可以发育成胚胎愈伤组织或没有胚柄的球形胚。甚至松散地围绕在原生质体周围的细胞壁似乎也起作用，并且与第一次合子分裂的方向和顶端 - 基部轴密切相关，这进一步表明原始合子细胞壁在维持顶端 - 基部极性和细胞分裂方向以及体外早期胚胎发育过程中随后的细胞分化方面的重要作用。
