【作者】 陈金慧；

【导师】 施季森；

【作者基本信息】 南京林业大学 ， 林木遗传育种， 2003， 博士

【摘要】 杂交鹅掌楸是一种重要的园林绿化和工业用材树种,杂交制种亲本资源有限,杂交种供不应求,通过细胞工程技术快速繁殖性状优良的再生植株,具有重要的应用价值和实践意义。 以杂交鹅掌楸幼胚为外植体,以植物细胞具有全能性的理论为依据,以细胞分化发育的分子调控理论为指导,进行体细胞胚胎发生研究。比较了亲本基因型、发育阶段、培养条件、生长调节物质和渗透压等对体细胞胚胎发生的影响。结果表明:在以1/2MS培养基为基本成分,添加2.0～3.0mg·L^-12,4-D的培养基上能够成功诱导胚性愈伤组织。以蔗糖为渗透剂,提高渗透压有利于体细胞胚胎发生;以MS为基本培养基附加ZT1.0～4.0mg/l对体细胞胚胎诱导最有效;ABA的使用能有效控制畸形胚的产生;基因型、幼胚发育阶段及光照条件对体胚发生起着重要作用。 通过组织细胞学方法研究杂交鹅掌楸体细胞胚的起源与发生部位,发现杂交鹅掌楸体细胞胚多是单细胞起源,表面发生,体细胞胚经过球形胚、心形胚、鱼雷胚和子叶胚而发育成再生植株,子叶期体细胞胚可观察到明显的V型维管束、茎尖分生组织以及根尖分生组织;以透射式扫描电镜和扫描电镜技术研究胚性愈伤组织与非胚性愈伤组织以及体细胞胚发育各个阶段扫面电镜下的形态差异、超微结构上的变化,发现胚性细胞结构较规则,细胞器相对丰富,细胞代谢旺盛;ATP酶的活性定位研究表明,胚性细胞的膜系统等部位存在活跃的ATP酶活性反应。 通过染色体计数对于再生植株的遗传稳定性进行研究,发现多数细胞系的再生植株遗传稳定性较好,为二倍体再生植株,仅在一个细胞无性系当中发现所有再生植株均为四倍体,说明我们建立的体细胞胚它发生体系遗传稳定性好。更多还原

【Abstract】 Somatic embryogenesis was successfully done in Liriodendron hybrids with the immature zygotic embryos as explants. Some factors had been tested about their effects on somatic embryogenesis in this research, such as genotype, developmental stage ,culture condition ,plant growth regulator and osmotic potential. The results showed that the embryogenic callus was successfully induced when the experimental materials were cultured on 1/2 medium supplemented with 2,4-D 2.0³.0 mg · L^-1. Sucrose can be used as osmoticant to improve osmotic potential of the medium and it was effective for somatic embryogenesis. The induction of somatic embryos was best when the embryonic callus was cultured on MS medium supplemented with 1.0 ⁴.0 mg · L^-1ZT. It was also proved that a proper amount of ABA can lessen the production of abnormal embryos. In addition, the genotype , developmental stage of immature embryos and light condition were important to somatic embryogenesis.Histological study on the origin and development of somatic embryos, it was found that somatic embryos were mostly developed from single embryogenic cells that possessed the ability of embryoid occurrence from the surface of the callus. At the early development stages , the proembryos were limited in a boundary of thick cell wall. Plantlet regenerations were developed from global embryo, heart-shaped embryo, torpedo-shaped embryo and cotyledon embryo, in which the V-shaped vessel vascular bundles and shoot tip meristem and root tip meristem were observed. By means of transmission scanning electron microscopy and scanning electron microscopy technique, it was found that the structure of embryogenic cells were regular and organelle were comparatively abundant . At the same time , we found that embryogenic cell metabolism were vigorous by studying the difference between embryogenic callus and nonembryogenic callus. ATPase activity was observed on the membrane system of the embryogenic cells.The genetic stability of the plantlets regeneration has been studied by counting the chromosomes and it was found that these plants were diploid mostly, while one of the clones and its regenerated plants all were tetraploid.更多还原