番茄叶组织培养中愈伤组织形成和器官发生的细胞学和微形态学研究

以番茄叶外植体为材料,研究了不同的生长素和细胞分裂素及其浓度配比对叶外植体培养行为的影响;同时,利用细胞学和扫描电子显微镜技术观察了愈伤组织形成和器官发生过程。结果表明,不同种类及浓度配比的生长素和细胞分裂素直接影响愈伤组织的物理状态、大小和形成的速度以及器官分化的频率和速度。叶外植体切口处的叶肉细胞,维管薄壁细胞和维管束上方的少数叶肉细胞首先启动脱分化而开始分裂,这些细胞的活跃分裂和分化导致在外植体表层形成由薄壁细胞、维管组织和无分化状态的表层分生细胞团组成的愈伤组织。而不定芽则通过愈伤组织的薄壁细胞再次脱分化和再分化活动而形成,为“外起源”。认为存在由植物激素决定的“无分化活性”和“有分化活性”二种性质的愈伤组织。     

椰子幼花序离体培养的器官发生

椰子幼花序在改良的ＭＳ附加高浓度的ＮＡＡ与２,４－Ｄ的培养基产生愈伤组织,在逐渐降离ＮＡＡ和２,４－Ｄ浓度的过程中,愈伤组织产生结构紧密的结节组织,结节组织有输导组织,分生细胞团,根原基及不同发育阶段的胚状体,有些进而发育成正常的根和绿色芽状物,同时,某些幼花枝的基部直接长出绿芽,经继续培养形成具有正常根,叶结构的完整植株,但茎尖无顶端分生组织,无叶原基,有的发育成花枝,甚至在顶部长出几朵发育不正     

沙枣(Elaeagnus angustifolia L.)胚状体的产生及激素对器官发生的作用

供试材料为野生种(Elaeagnus angu-stifolia L.)的子房、茎和叶。在附加0.5毫克/升NAA和1毫克/升玉米素或再加2毫克/升2,4-D的MS培养基中均能诱导出愈伤组织。愈伤组织转入MS+KT(2毫克/升)或KT(2毫克/升)+IBA(0.2毫克/升)的分化培养基上可分化出大量的绿苗。苗分正常苗和异常苗两种类型,目前为止,沙枣愈伤组织无性系已继代培养了22代,将近两年的时间仍具有较强的分化能力。外源激素对正常苗的分化有直接的影响,高浓度的KT水平可促进正常苗的分化,其最佳浓度为4毫克/升,正常苗诱导频率可高达83—86%,基本上解决了分化的问题;高浓度的6-BA虽也能提高分化率,但异常苗占优势。试管苗形成的途径有二,一是通过不定芽的方式产生,一是通过胚状体的方式产生。沙枣胚状体原始细胞的来源有三种情况:(一)由紧邻表皮细胞的单个薄壁细胞产生,这种方式占优势;(二)由表皮细胞横裂产生;(三)由表皮细胞及其下面相邻的薄壁细胞同时分裂共同参与胚状体的形成,即二者的嵌合体。胚状体的发生与合子胚的发生过程基本相似,但在其发育的早期阶段无典型的基细胞与顶细胞之分,故也缺乏典型的胚柄结构,随着胚状体的长大,表皮细胞被撑破,其周围的薄壁细胞内含物被吸收而解体,最后胚状体脱离开愈伤组织的表面,孤立出来成一个完整的个体。     

蒲圻贝母组织培养鳞茎形成过程中生理生化变化研究

对蒲圻贝母鳞茎的组织培养进行了研究,筛选出了较佳的激素配比,并对鳞茎生长过程中的可溶性糖、淀粉含量、淀粉酶活性、ＰＡＬ酶活性以及生物碱含量变化之间的关系进行了探讨。     

COMPARATIVE ANATOMY OF ENDOGENOUS BUD AND LATERAL ROOT FORMATION IN CONVOLVULUS ARVENSIS ROOTS CULTURED IN VITRO

The anatomy of endogenous bud and lateral root formation was studied in Convolvulus arvensis roots cultured in vitro. Although early stages in the initiation of buds and roots were found to be identical, continued development of primordia into roots or buds was accompanied by differences in participating primary root tissues, in rates of development, and in orientation of cell divisions. Evidence for the existence of a primordium capable of developing into either a bud or root is discussed with reference to Convolvulus roots and other plant parts shown to have similar morphogenetic potentials.     

GROWTH RESPONSES FROM WHOLE FRUIT AND FRUIT HALVES OF LEMON CULTURED IN VITRO

A comparative growth study was conducted on juice vesicles cultured in the form of various fruit explant types (equatorially bissected fruit halves, longitudinally bissected fruit halves, one-eighth sections of fruit, one-quarter sections of fruit, whole carpel segments, 2 or 3 mm thick equatorial slices of fruit, and 1 cm² fruit endocarp pieces) from 15 mm diam Citrus limon (L.) Burm. f. cv. Eureka lemons. Juice vesicles within equatorial fruit halves produced the least amount of callus. Furthermore, these juice vesicles grew similarly to juice vesicles occurring in the tree grown fruit. A study of cultured equatorial fruit halves using 10–45 mm diam lemons was then conducted. Fruit half cultures containing juice vesicles could be readily established from 15–45 mm diam lemons. Vesicles from 10 mm diam fruit halves, however, invariably produced callus. Vesicles cultured within fruit halves produced proportionately less callus as their fruit diam increased. Juice vesicles cultured in 15–30 mm diam fruits lost their original green color and turned opaque as they matured (i.e., after 3–6 months in culture). A method is also presented, whereby whole lemon fruits can be established and maintained in vitro. Lemons, 35–45 mm in diam, were the best explant sources for establishing whole fruit cultures. Juice vesicles in whole fruit cultures may remain viable for up to 8 months in culture.     

INVESTIGATIONS ON THE ROOT FORMATION IN THE TISSUES OF HELIANTHUS TUBEROSUS CULTURED IN VITRO

Excised rhizome fragments of Helianthus tuberosus ‘Violet de Rennes’ produce roots in vitro; the sequence of morphogenetic phenomena is as follows: proliferation, differentiation of phloem and tracheids, organization of cambiums, and formation of root primordia from cambiums. This rhizogenesis is conditioned by five limiting factors: mineral salts, sugar, auxin, temperature, and light. The optimum level of each of these factors was determined and in certain cases also the threshold of action. For white light the beginning was about 1/1000 lux. These five limiting factors do not represent the same site of action. The mineral salts, especially nitrogen and calcium, have their effect when the cambiums have been organized. The sugars act before the formation of cambiums. Light operates principally upon cambium and this action is independent of photosynthesis. It was not possible to determine whether the auxin has a preferential site of action. A temperature higher than 15 C is essential to obtain the formation of cambiums but further production of roots is possible at 15 C or even less. The variations in temperature more or less around the optimum stimulate rhizogenesis. The supra-optimal temperatures mainly have their effect before the formation of cambiums and infra-optimal temperatures after. The effect of light can be exhibited at low temperature. Gibberellic acid in association with auxin favors root formation in the dark whereas in light it exhibits inhibiting properties. Rhizogenetic potentialities of tissues are reduced when these are cultured in the dark in presence of auxin. On the other hand, if auxin is applied after the isolation of explants, rhizogenesis is stimulated and the tissues are able to produce roots even in the dark. The action of light and of auxin can be dissociated; when the explants having absorbed a small quantity of auxin are transferred to a medium without auxin for 60 days in the dark, they conserve the rhizogenetic potential which is expressible on transferring them to light. Finally, grafting experiments suggested that light could induce the formation of a rhizogenetic factor in the tissues which is transmittable from cell to cell. These experiments have definitely established that the induction of root formation is governed by several factors which have complex interactions.     

CALLUS FORMATION AND ORGANOGENESIS FROM CULTURED LEAF SEGMENTS OF CRASSULA ARGENTEA: CYTOKININ-INDUCED DEVELOPMENTAL PATTERN CHANGES

Culturing leaf segments on agar medium with a final concentration of 10^–4m isopentenyl adenine inhibits regeneration of plantlets, but stimulates some segments to produce regions of dense callus. The callus is initiated from the basipetal end of a cut vascular bundle, and is therefore the positional equivalent of root initiation during normal regeneration. Once callus has emerged, it forms a sphere of tissue attached to the leaf segment. The cells on the surface of the sphere are smaller than the cells toward the center. Some of these larger cells contain tannins or large starch grains. Shoots are sometimes initiated from organized meristematic regions of surface cells. Roots form endogenously from callus after shoot formation. This regeneration sequence, initially induced by cytokinin treatment, is strikingly dissimilar to the normal pattern, and provides an excellent example of a hormone-induced alteration to a developmental pattern.     

六种中药及其复方对鳗鲡致病性气单胞菌的体外抑制作用

针对病鳗内脏中分离的5 株致病性气单胞菌, 采用琼脂稀释法测定五倍子、石榴皮、大黄、虎杖、黄芩及黄连各味中药的最低抑菌浓度(MIC)和最低杀菌浓度(MBC); 再根据棋盘法设计15 种双联用药方和4 种三联用药方, 同样检测各组合配方的抑菌作用。实验结果表明, 6 味中药对养殖鳗鲡5 株致病性气单胞菌均有不同程度的抗菌效果, 其中五倍子的抑菌作用最强, 其次是大黄和石榴皮, 而黄连的抑菌效果最差; 15 种双联用药方较各味中药单用的抑菌活性绝大多数出现增强, 抑菌浓度至少减低39%, FICFIC≤0.5 表现显著增强抗菌活性的协同比例占23.3%; 4 种三联用药方对5 株致病性气单胞菌均具有显著的协同抑制效应, 复方中单味中药的抑菌浓度可以降低80%以上; 而双联用HC14 对4 株致病菌出现FIC≥2 的降低彼此抗菌活性的相互拮抗现象。由此说明合理运用不同中药的联用配伍, 不仅可提高单味中药的抗菌疗效, 而且大大减少了单一中药在实际养殖生产中的给药浓度, 降低药物在环境中的残留量, 防止残留药物造成环境污染, 并且降低用药成本, 提高水产养殖业的经济和社会效益。研究为中药复方防治细菌性鱼病提供科学理论参考。       

宁夏枸杞器官发生和体细胞胚胎发生过程中DNA、RNA和蛋白质合成动态的比较研究

宁夏枸杞(Lycium barbarum L.)叶外植体来源的愈伤组织经筛选、繁殖后,将来源相同、状态较为一致的淡黄色愈伤组织转移至O型或E型培养基上,可以诱导出器官发生和体细胞胚胎发生。利用该体系,对两条离体再生途径进行了比较研究。结果表明:(1)在拟分生组织和胚性细胞形成之前,RNA合成首先被激活,随后DNA、蛋白质合成加速;而球形胚形成期间,先是DNA合成的加快,接着RNA、蛋白质的合成高峰出现,在不定芽形成期间却正好相反;(2)可溶性蛋白组分发生规律性变化;器官发生和体细胞胚胎发生的启动阶段都有-153.6kD多肽出现,一些多肽分子在分化早期逐渐消失,而随芽原基或球形胚的形成又重新合成;与形态发生相对应,两种再生体系都有作为各自分子标记的特异多肽(84.9kD、46.3kD和44kD、36.2kD)的表达。此外,还对两种离体再生体系之间的关系和发生机制进行了讨论。     

THE HORMONAL CONTROL OF ORGAN FORMATION IN CALLUS OF MEDICAGO SATIVA L. CULTURED IN VITRO

Organogenesis in alfalfa callus (Medicago sativa L. cv. ‘Regen S’) has been obtained by the transfer of callus from an induction medium containing growth regulators to a regeneration medium lacking growth regulators. The transfer of callus from induction medium containing high levels of 2,4-D and low levels of kinetin to regeneration medium resulted in the formation of shoots. Conversely, the transfer of callus from induction medium containing low levels of 2,4-D and high levels of kinetin resulted in the formation of roots. The pattern of organogenesis on regeneration medium was modified by the nutritional composition of that medium. When Blaydes medium supplemented with inositol and yeast extract was employed as regeneration medium, root organogenesis was inhibited. Root organogenesis was not inhibited by either Shenk and Hildebrandt medium or Gamborg's B5 medium. Shoot formation occurred on all of these media. A survey of the in vitro organ-forming capacity of 14 genotypic clones from the cv. ‘Regen S’ was conducted. The capacity to form organs differed quantitatively among the clones analyzed. A more detailed analysis of a highly responsive clone (RA3) and a poorly responsive clone (RA5) revealed no significant qualitative difference in their organogenic responses.     

青霉素和NAA与6—BA配合在离体草莓器官建成中的作用

本实验研究了青霉素和ＮＡＡ与６－ＢＡ配合对离体草莓器官建成以及体内过氧化物同工酶的影响。结果表明：青霉素促进不定根的分化,但抑制不定根的伸长和不定芽的分化,并且不同程度地影响器官建成过程中过氧化和抽工酶的谱带和活性。青霉素与植物激素一样,参与了植物体内的生理代谢而引起植物器官建成。     

新合成的蛋白质和RNA从离体萌发的花粉中释放

应用同位素放射自显影技术,以~3H-尿嘧啶和~3H-亮氨酸作前体,对人工萌发花粉中新合成的 RNA 和蛋白质进行标记;并借助 RNA 合成抑制剂放线菌素 D 和蛋白质合成抑制剂环己亚胺的对比试验,确证了君子兰花粉在萌发过程中,向外释放了新合成的 RNA 和蛋白质。动态如下:标记后1小时,花粉尚未萌发,大量的 RNA 和蛋白质已合成;2小时后,新合成的蛋白质向花粉管方向转移;5小时后扩散到培养基中。新合成的 RNA 在90分钟内暂留在营养核和生殖细胞中;2小时后扩散到花粉细胞质;3小时后集中到花粉管顶端;5小时后释放到花粉外。文章还讨论了技术的关键问题和现象的生物学意义。     

水稻游离花粉培养的高频率再生植株

用二个水稻栽培品种(Oryza sativa L Sub.japonica.)中花11号和盐粳的花粉处于单核靠边期的花药,经低温处理10—20天,在无糖培养基中预培养2—4天后游离花粉进行培养。培养基为KM8P,附加1mg/L 2,4-D,100mg/L脯氨酸,500mg/L水解酪蛋白,9%蔗糖。培养5天,花粉进行一次分裂,10天后分裂频率为21.3%,21天可见小愈伤组织形成。随即将直径为0.5—1.0mm的愈伤组织移至分化培养基上,2—4星期后得到绿苗,频率为70%。并从许多花粉诱导的愈伤组织克隆中筛选出高频率再生绿苗的花粉细胞悬浮系。     

五味子甲素对大鼠成骨细胞增殖分化的影响

目的　探讨五味子甲素体外对大鼠成骨细胞增殖与分化的影响。方法　用改良的组织块法分离培养新生大鼠颅骨成骨细胞 ,五味子甲素以不同浓度加入细胞培养体系 ,作用不同时间后 ,用MTT法检测成骨细胞的增殖情况 ;用对硝基苯二钠基质动力学法 (PNPP)测定细胞内碱性磷酸酶的活性 ,用改良的Lowry法测蛋白含量。 结果　五味子甲素在 0 75× 10 -5mol/L及 0 75× 10 -9mol/L浓度范围内 2 4h ,72h ,0 75× 10 -4 ～ 0 75× 10 -7mol/L 48h促进成骨细胞增殖 ,在 0 75× 10 -5～ 0 75× 10 -6mol/L浓度范围内 48h提高成骨细胞内碱性磷酸酶的活性。结论　五味子甲素体外能促进成骨细胞的增殖与分化。     

蛇床子素对大鼠成骨细胞增殖分化的影响

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槐树试管正常苗与超度含水态苗茎叶的比较形态学研究

槐树(Sophora japonica L.)子叶经培养获得了大量的试管苗,依其形态正常与否可将其分为正常苗、超度含水态苗和介于二者之间的过度含水态苗。其自由水含量明显不同,正常苗低于50％,超度含水态苗高于79％,而过度含水态苗约为70％。以扫描电镜对其茎、叶的形态学结构进行了比较研究,结果表明:正常苗茎、叶表皮结构基本类似于实生苗,而过度及超度含水态苗的茎、叶表皮层结构变异较大。主要表现在其表面凹凸不平,表皮层外稀有或无蜡质存在,表皮细胞形状及排列不规则,叶片近、远轴两面气孔器密度、大小及开度均较正常苗显著增大;保卫细胞形态、结构异常。上述特征,均显示出槐树试管过度及超度含水态苗易失水干化,这可能是其在移栽过程中难以成活的主要原因之一。