【作者】 冯金玲；

【导师】 陈辉；

【作者基本信息】 福建农林大学 ， 森林培育， 2004， 硕士

【摘要】 锥栗的经济价值很高，是一种粮材两用的树种，树体各个部位都具经济价值，如木材可以做家具，花是蜜源，叶子是蚕食等，特别果实是一种不可多得的木本粮食，果肉中含有大量的淀粉、蛋白质和各种维生素。但锥栗是扦插不易生根的植物。目前，锥栗的品种混乱，病害严重，且对锥栗的研究大部分是宏观的，微观上的研究相对少。 本研究通过组织培养的手段，建立锥栗的再生体系，为锥栗良种选育，定向培育及遗传转化和基因克隆等方面的研究，提供基础手段和理论依据。 本研究采用的材料来自于福建省建瓯市水源乡桃源村，分别把油筒子、油榛、黄榛、大尖嘴、油栗子、白露子、中尖嘴、乌榛、乌壳长芒、欧宁子、坝头子、处署红、穗榛这13个品种的不同部位作为外植体，如胚、腋芽、韧皮部等，进行愈伤组织培养、增植愈伤组织培养、分化愈伤组织培养、诱发不定芽培养、增植不定芽培养及生根培养，形成再生植株，移栽成苗。研究了基本培养基，激素种类和浓度，品种，外植体类型，接种方式，褐化，糖的种类和浓度，维生素B₁，赤霉素，培养基的其它有机添加物，琼脂，pH和外界环境条件等因素对锥栗组织培养的影响。得到了如下结果： 1．升汞和次氯酸钠不同浓度在不同时间下对锥栗胚、腋芽、韧皮部等外植体进行消毒，其效果差。但在75％酒精消毒果仁的情况下，直接取的胚可以很好的控制污染率。 2．13个品种的锥栗作为材料，得出最佳品种是油榛，其次黄榛、大尖嘴等，接着是处署红等，而最差等级是油筒子、穗榛、白露子等。锥栗的不同器官作外植体时是以胚为最好。并且以胚根插入培养基为最好的接种方式 3．锥栗愈伤组织培养途径的基本培养基是MS，在诱导愈伤组织时生长素2，4-D优于NAA和IBA。愈伤组织增植时生长素优于分裂素，而且在生长素中以2，4-D最好。对于愈伤组织的分化，虽然整体上分化不理想，但实验也得到一些结果：分裂素BA对锥栗分化的效果优于KT，生长素IBA优于IAA，对于分裂素BA，随着浓度从低到高，其分化效果从好到差；当分裂素BA为5mg／l时，愈伤组织的生长受到威胁，甚至褐死；在分裂素与生长素的作用效果上看，当BA为3mg／l时NAA为0.5mg／l时分化效果好。当BA为1.5mg／L时，NAA0.2mg／l和NAA0.5mg／L为佳，可见不同种类浓度的分裂素与之搭配的生长素的浓度和种类也不同；在AgNO₃对愈伤组织的作用中，AgNO₃ 1mg／l对锥栗愈伤组织的分化没有起到作用。 4．锥栗不定芽生长的基本培养基是M基本培养基。在锥栗不定芽的预实验中得到了锥栗不定芽诱发的BA以低浓度1mg／L好于其它的高浓度的锥栗组织培养及其再生体系建立的研究 处理，在生长素对锥栗不定芽诱发的实验中得到份A对于锥栗的芽生长 是最适合的，其次是NAA，最后是IAA。激素配比的实验中，锥栗的 胚在分裂素与生长素的比例为8:l时，锥栗能诱发出正常的不定芽。 但并不是相对含量越高或者越低越好，而是有一定的适应范围，得出三 种好的处理，分别为BA十NAA二Zm幼十0.2一0.4m酬， BA+IBA=2m幼+0，4nlg/1，BA+IBA“1 .sm幼+0.2mg/l。且以 BA+I BA司.sm酬+0 .2m酬处理的激素配比为最佳。锥栗不定芽增植的最 佳激素配比是BA+认A=o.75mg/l+o.4m酬。5.锥栗不定芽促根的基本培养基是M基本培养基。在激素的含量与种类 对锥栗不定芽生根的影响实验中生长素IBAlm留1对锥栗的不定芽促根 起到极显著作用。6在锥栗抗褐变的实验中，添加不同的抗褐变的物质，其结果只有AC起 到比较明显的作用，而PVP和N匆503只发挥较小作用，但抗坏血酸却 起反作用。在锥栗碳源的种类和浓度实验中，得到蔗糖是最佳的碳源， 且最佳的蔗糖浓度为40%。在维生素B，的实验中得到维生素Bllm留l 对锥栗的不定芽生长有促进作用。在组织培养中加入一些有机营养成 分，如水解酪蛋白（HC），谷氨酞氨，叶酸（维生素BC）和D一泛酸钙， 对锥栗的不定芽生长起到了抑制作用。GA对锥栗的不定芽有促进作用。 在琼脂浓度对比实验中，锥栗外植体生长最适合的浓度是7%。锥栗培 养物生长的最佳pH值是5.5。在N氮素对比实验中，NH3No;和KNO3 的最佳含量分别为825m酬和950m幼。Fe因子对比实验得到最好的Fe 含量是27.smg/l。7.本实验对光照的强度和温度分别作研究，并得出锥栗最适合光照是 20001ux光强和最适合温度是28oC。8.锥栗的植株生出了完整的根系后，可进行移栽。进行炼苗，先在光照培 养室中旋开瓶盖，静放2一3天，然后再把带苗的瓶子拿到自然光的房间 中放4天左右，最后把带苗的瓶子拿到强自然光的外界环境中放2天左 右。然后把苗移栽到沙基质中。更多还原

【Abstract】 Castanea henryi , both of its seed and wood being useful , has very high economic worth , for example, its fruit tasted well and enjoyed for a long time, its timber for making furniture, its flower as nectar source, etc.. Its fruit is a kind of rare woody grain, contain a large number of starch , protein and various kinds of vitamin in the pulp. But the cloning is uneasy to make Castanea henryi rooting. At present, there are many problems in the research of Castanea henryi, such as the confusing cultivar series , the serious disease, etc., and some studys on Castanea henryi are nearly in the macroscopic field , little in the microcosmics field.This research, for setting up the regeneration system of Castanea henryi by means of tissue culture offer the basic means and theoretical foundation for stock breeding, orientation fosters and heredity transform and gene cloning, etc..The material adopted in this research came from Shuiyuang village of Jianou town in Fujian Province. There are 13 cultivars of Castanea henryi, such as Youtongzi, Youzhen, HuangZhen, DaJianZhui, YouLiZi, BaiLuZi, ZhongJianZhui ,WuZhen, WuKeChangMang, OuNingZi ,ChuSuHong, ShuiZhen, from which the embryo, axillary bud, bast etc. were taken as explants for being cultured and regenerated into plantlets, by means of callus culturing, enrich callus culturing , differentiation callus culturing, induced adventitious bud culturing, enrich adventitious bud culturing and rooting culturing . Various elements and hormones, hormones concentrations, varietys of explants, the inoculation way the browning sucrose kinds and concentrations the vitamin B1. the gibberellin and other organic nutrition in culture medium, such as the agar, the pH and the external environmental condition etc., were tested in tissue culture of Castanea henryi. The following results had been received:.1. The different concentrations of mercuric chloride and sodium hypochlorite disinfected to the explants of embryo , axillary bud, basts under different time, worked not well. But the embryos which directly draw from the kernels disinfected by 75% of the alcohol are better to controlling pollution rate.2. Taking care of 13 cultivars of Castanea henryi as material , YouZhen is regarded as first-class ones, HuangZhen and DaJianZhui etc. as second, ChuSuHong etc. as third, YouTongZi, ShuiZheng and BaiLuZi etc. as the fourth. When different organs of Castanea henryi were taken as explants, the embryo was regarded as the best. In inoculation way, the radicle insertingculturing was regarded as the best.3. The basic culture medium of callus culture is MS. When callus was induced , 2,4-D was better than NAA and IBA. while enriching callus, the growth hormone is better than the mitosin, and 2,4-D is the best kind of the growth hormone. Differentiating callus showed that BA is superior to KT, IBA is superior to IAA. The differentiating effect changed from good state to bad one as the decrease of the density of B A. The callus had hardly vitality and was likely browning to death when the concentration of BA reach to 5 mg/1. Considering the function of mitosin and the growth hormone , when the 3 mg/1 of BA matched with the 0.5 mg/1 of NAA and the 1.5 mg/1 of BA matched with the 0.2 mg/1 of NAA0.1 or the 0.5 mg/1 of NAA ,the result of differentiation was good. It is obvious that different kinds of density mitosin matched with the different kinds of density growth hormone. AgNO3 1mg/1 had not function on the callus defferentiation.4. The basic culture medium of Castanea henryfs adventitious bud is M medium. In preliminary test of induced adventitious bud, BA with low density as 11.5mg/l was better than other higher density treatment. IBA is the best kind of growth hormone, the second is NAA, and the third is IAA. In the experiment of hormone matching, when the proportion of the mitosin and the growth hormone reached 8:1, the normal adventitious bud could be brought out. But there were certain accommodations, and the experiment tested out three kinds of good treatments: BA+NAA=2m更多还原