||甘草自古就是一种重要的药用植物,同时在食品、化妆品、烟草、轻工、石油、消防等许多行业中有非常重要的作用,且又是我国西部地区重要的固沙植物.近年来,甘草需求量不断增加,野生甘草被过度采挖并呈现濒危状态,人工种植甘草逐渐成为甘草研究的热点,选育品质优良的甘草品种是加大人工甘草推广种植面积、缓解野生甘草危机的唯一出路.植物的授粉方式直接影响到植物本身的遗传特点、育种策略、种子生产方法和栽培管理技术等,植物胚胎学是进行植物遗传育种研究的基础.鉴于目前甘草授粉生物学特性缺乏统一认识,不能有效的指导育种实践,本研究以内蒙克什克腾旗和新疆福海两地的野生甘草为实验材料,通过花器形态学观察、访花昆虫观察、杂交指数和花粉-胚珠比测定、田间授粉实验、授粉前后雌雄配子发育研究以及AFLP分子标记等方法,研究了甘草的有性繁育系统,主要研究结果如下:1甘草花器的形态学特点表明甘草具有异花授粉的特征:甘草为总状花序,垂直型,自下而上顺序开花.小花较大,在10-20 mm之间,花冠自从花萼中露出开始至凋谢均为鲜艳的紫色或浅紫色,花粉粒近球形,表面具有疵状或颗粒状突起或无,在开花初期,甘草柱头高于花药,二者存在1-1.5mm的空间距离,至开花后期大约有20%的柱头高度和花药持平.该结果表明同花花粉到达柱头的几率很小,很难进行自花授粉,具有异花授粉的特征.2甘草访花昆虫种类较多:主要有蜂类、蝶类、蝇类、蚂蚁、蚜虫等,蜂类在花朵上停留时间20-30秒,蝇类停留时间长,在3-4分钟,二者均具有明显的授粉行为.蚂蚁和蚜虫在花冠、柱头和花粉上的爬行是间接的传粉行为.3甘草花粉生活力在散粉当天最高,柱头最佳可授期为开花后第二天:本实验在散粉前后进行花粉生活力测定实验表明甘草花粉在散粉当日的花粉活力最高,为94%,散粉后第2d和第4d,花粉活力有所下降,但都维持在90%以上,之后则迅速下降,到第16d,花粉全部死亡,活力完全丧失.甘草柱头活力与花的发育时间有关,其柱头的寿命为3d,柱头在小花开花后第2d活力最高,为授粉的最佳时间.4甘草杂交指数和花粉-胚珠比表明甘草繁育系统为兼性异交类型:甘草花朵直径大于6 mm,雌蕊先熟,柱头和花药有1-1.5mm的空间距离,杂交指数OCI为4,表明甘草的繁育系统为异交,部分自交亲和,需要传粉者.甘草每朵小花平均花粉量为3600粒,每朵小花平均胚珠数为10个,P/O在309~429之间,表明繁育系统为兼性异交类型.5不同方式的授粉处理表明甘草自交亲和,异株授粉结实率更高,为兼性异交类型植物:通过对内蒙和新疆两地的甘草进行套单花、套单株花序、人工同花序异花授粉、人工同株异花序授粉、人工异株授粉实验,结果甘草在自然条件下的结荚率仅为10%,套单花的处理没有结实,套花序处理结荚率仅为1.6%,但人工控制授粉的处理结荚率明显提高,人工同序异花授粉的结荚率为14%,人工同株异花授粉的结荚率为25%,人工异株异花授粉的结荚率为58%,表明甘草自交亲和,但缺乏自花传粉媒介,人工异株授粉结实率更高,为兼性异交类型植物.6 AFLP分子标记明确甘草基因型具有一定的杂合性:分别选择来自哈密、阿拉尔、和田、巴楚和莎车的5个亲本各50个后代植株,利用筛选的9对引物对总DNA进行扩增片段长度多态性AFLP分析,发现来自同一亲本的不同后代单株的差异性条带百分率为19.32%,表明甘草的基因型具有一定的杂合性而不是完全的纯合体.7甘草授粉前后胚胎学发育特点表明受精后正常发育胚珠数量少是甘草结实率低的主要原因:本实验以现蕾后不同发育阶段的小花为实验材料,明确了甘草花药在减数分裂后小孢子为四分体型,胚珠为弯生胚珠,受精时先经过两个极合的靠近或融合后再与精细胞融合.受精后正常发育的胚珠数量少是造成甘草结实率低的原因之一.
||Glycyrriza Uralensis Fisch is an important medicinal plants since ancient times, and also plays a very significant role in many industries such as food, cosmetics, tobacco, light industry, petroleum and fire system. Meanwhile Licorice is the important sand-fixation plant in western areas of China. Recently, with the increasing demand of Licorice, the wild Licorice has been over collected and endangered, while the artificial cultivation has been the focal topics in the research of Licorice. To select high quality strain of Licorice is the only option to enlarge the planting areas of the artificial cultivation and alleviate the crisis of the wild Licorice.The pollination method of plants can directly affects genetic characters, breeding strategy,seed production methods and cultivated management technology. Plant embryology is the fundamental to study the genetics and breeding of plants. Considering lack of a consistent understanding of the technique of artificial-pollination of the Licorice, which was unable to guide the breeding practices, this study used the wild Licorice for the experimental materials from Keshiketeng county, Inner Mongol Autonomous Region and Fuhai in Xinjiang, studied the biological pollination of the Licorice by the observation of floral organs' morphology, the experiment of field pollination, the experiment of pollen tube's germination labeled with fluorescent markers, the study of the development of male and female gametophyte around the pollination as well as the molecular markers of AFLP.1. The morphological characteristics of the Licorice's pollen tube suggests that the Licorice has the character of Cross-pollination: The little flowers of Licorice are larger, the variation ranges from 10 to 20mm,and the corolla is bright purple or pale purple firstly blooming out of calyx. Pollen grains are stereo and in near spherical shape, and the surface has granular apophysises or not; the space distance between stigma and anther is 1-1.5mm, and the pollen can hardly reach the stigma; while the Licorice has vertical inflorescence, and the sequence of bloom is from bottom to top. Thus, from the characteristics of the external morphology of the Licorice, it has the character of cross-pollination.2. The experiment of field pollination preliminary judge that the Licorice is a plant of Cross-pollination:By experimenting on the Licorice from Inner Mongolia and Xinjiang with the single flower covered, single plant's inflorescence covered, same inflorescence be cross pollination artificially, monoecious be pollinated by different inflorescence, cross pollination among the individuals, the result was that the pod rate of Licorice in natural condition was only 10%..The treatment with single flower covered has no pod, and the pod rate of single plant's inflorescence covered was only 1.6%,but the pod rate of artificial pollination improved significantly, the pod rate of same inflorescence pollinated by different flowers was 14%,same plant pollinated by different flowers was 25%,while different plants pollinated by different flowers was 58%. It suggests that Licorice is a Cross-pollination plant, and artificial pollination can improve the self-fruitful rates of Licorice. Also, the pollinators of Licorice are bees by tracing observation of camera.3. The molecular markers of AFLP determined the genes of Licorice were heterozygous:Choosing five parent each of 50 plants, respectively from Hami, Alar, Hotan, Bachu and Yarkand, and using screened 9 primers to make analysis of AFLP for total DNA, discovering that the percent of difference bands from the same parent's different offspring's plants is 19.32%.It suggested that the genotypes of Licorice were heterozygous but not exactly homozygote.4. The germination rates and the length of elongation of Licorice's pollen tubes under different pollination methods are not alike. it suggests that after the pollination, the pollen disrupting rate was up to 15-20%, the germination rate of pollen tube after pollination can be 30%, but the length of elongation of pollen tube was short that unable to reach the ovules to complete the double fertilization; suggesting that the short germination of pollen tube after pollination is one of the reasons for self-incompatibility of Licorice. While pollen tubes' germination rate after the artificial cross pollination between different plants was up to above 90%, and germinated quicker, so it can reach the ovules quickly to complete the fertilization and develop vigorous seeds. This test supports the conclusion that Licorice is a cross pollination plant.5. The pollen viability of the licorice is the highest on the day of anther dehiscence, the best pollination period of stigma is the second day after flowering. The samples were collected periodically around the anther dehiscence, to undergo TTC staining to determine the pollen viability. The result shows that, on the day of anther dehiscence, pollen viability is up to 94%, the 2nd and