棉花组织培养与植株再生

棉花是世界上重要的纤维和油料作物。现代生物技术的不断发展为棉花育种和种质创新提供了新的技术手段,极大地推动了棉花育种的进程。现代生物技术在植物育种中的成功应用大多是依靠有效的再生体系的建立。因此棉花组织培养及其再生技术在品种改良、种质资源的繁殖和保存、杂种优势的固定和遗传转化等方面具有重要作用,颇受各国研究者的重视。本文综述了棉花的体细胞培养、花药培养、茎尖培养、胚珠和胚培养以及原生质体培养等方面近年来的主要研究进展,并提出了棉花组织培养中存在的主要问题及今后的研究方向。     

植物原生质体分离及培养研究进展

原生质体分离及再生体系现已被广泛用于生理、生化、遗传、分子生物学、基因组学、蛋白质组学、代谢组学的研究中,并能够为植物遗传转化提供良好的受体系统,从而达到优化遗传性状,丰富种质资源的目的等。园艺植物由于其观赏性强、倍性高的特点,具有很高的研究价值,但目前具有稳定的原生质体分离体系的园艺植物数量很少。本研究总结了园艺植物原生质体分离及培养时的各种影响因素：原生质体分离的植物材料;原生质体的酶解条件和纯化;原生质体的培养方法;原生质体的培养基成分,以及原生质体在各领域的应用,为建立更多植物原生质体分离及再生体系提供参考,为将原生质体系统更广泛的应用于多种植物中提供帮助。     

棉花黄萎病菌培养滤液对棉幼苗、种子发芽及离体培养物的毒害作用

用试验证实了Ｃｚａｐｅｋ＇ｓ液体培养基培养棉花黄萎病菌所得病菌培养滤液，能使棉苗发病萎蔫甚至干枯死亡；测试了病菌培养滤液对种子发芽、根的生长、愈伤组织诱导、愈伤组织生长及原生质体活力的影响，抗感不同品种表现出较明显的差异。为病菌培养滤液应用于抗病资源的快速筛选与鉴定、体细胞抗性突变体的筛选等研究提供了可靠的参考资料。     

二倍体棉种组织培养研究进展

二倍体棉种为四倍体栽培种提供了亲本(祖先)种,是棉花遗传改良的重要种质资源,是构建棉属突变体库、研究棉花功能基因组的优良材料。其中A、B、E、F组棉种起源于亚非大陆,D组起源于美洲大陆,C、G、K组起源于澳洲等地,具有丰富的遗传多样性,其组织培养研究主要涉及植株器官与细胞培养的再生途径、原生质体培养和体细胞培养与融合等,目前已获得了克劳茨基棉(Gossypium klotzschianum)、野生拟似棉(G. gossypioides)、亚洲棉(G. arboreum)、戴维逊氏棉(G. davidsonii)、雷蒙德氏棉(G. raimondii)、司笃克氏棉(G. stocksii)、奈尔逊氏棉(G. nelsonii)、比克氏棉(G. bickii)等二倍体棉种的再生植株,其中茎尖培养仅获得了亚洲棉、比克氏棉再生植株;原生质体培养获得了克劳茨基棉、戴维逊氏棉的再生小植株;此外,获得了陆地棉和二倍体棉种克劳茨基棉、比克棉、司笃克氏棉的种间杂种。二倍体棉种存在着较大的种间差异,组织培养体系可以借鉴木本植物组织培养相关方法。本文综述了相关研究进展,讨论了后续研究方向。     

原生质体技术与观赏植物育种

原生质体培养和植株再生技术在观赏植物育种中起重要作用。原生质体的来源、原生质体的分离、培养基的成分、培养方法等,都对原生质体的培养有重要影响。近年来,观赏植物原生质体在细胞融合、遗传转化上有了一定发展,为创造观赏植物新种质提供了新途径和方法。     

果树原生质体培养研究进展

原生质体培养是进行植物育种和细胞各种生理生化特性研究的平台.本文对近些年在果树原生质体分离培养条件、方法和植株再生技术方面的研究进展进行了综述,分析了影响原生质体的分离、纯化、培养的有关因素,并根据有关文献讨论了今后果树原生质体研究的重点方向.     

棉花组织培养体细胞胚胎发生的扫描电镜观察

近年来棉花组织培养发展很快, 先后从陆地棉、 海岛棉、 克劳茨基棉、 戴维逊氏棉、 拟似棉等不同棉种的胚轴、 子叶、 茎段、 花药、 原生质体等诱导获得了体细胞胚胎发生和植株再生。 但是对棉花体细胞胚胎发生的机制还需要探索, 本文采用扫描电镜技术对棉花体细胞胚胎发生的形成过程和形态特征进行了研究, 旨在进一步了     

科技新闻

棉花原生质体培养再生植株获得成功最近山西省农科院与江苏省农科院合作,成功地从棉花胚性细胞原生质体中培养出再生植株。这一结果迄今未见报道。至此,我国主要农作物—水稻、小麦、玉米、大豆、棉花的原生质体培养再生植株均已获得成功。……     

水稻与空心莲子草原生质体电融合条件的研究

以水稻（Oryza sativa L.）的胚性悬浮细胞和空心莲子草[Alternanthera philoxeroides(Mart.)Griseb]的悬浮细胞为材料， 研究了两种原生质体电融合的条件。结果表明：在交流电场为100V/cm，直流脉冲电场为1.5kV/cm，脉冲幅宽为40μs，脉冲间隔为0.5a，脉冲次数为2次，电融合液组成为10％甘露醇和0.4mmol/LCaCl2时，原生质体的融合频率达到24.6％。     

原生质体载体技术在马铃薯育种中的应用

综述了原生质体载体技术在育种中的应用:体细胞杂交、以原生质体为受体细胞的DNA直接导入转化、体细胞无性系变异、胁迫耐受系的选择,同时针对马铃薯的生物学特性、育种现状及其原生质体方面的研究现状,提出利用原生质体载体技术进行马铃薯育种的优势和切实可行性。     

Production of somatic hybrid plants between Iris ensata Thunb. and I. germanica

Wide hybridization that cannot be attained through conventional sexual crosses, can now be approached by somatic hybridization. Protoplasts of I. ensata and I. germanica were fused by electrofusion. For the selection of somatic hybrids, protoplasts of I. ensata which did not form colonies in protoplast culture and protoplasts of I. germanica which had regeneration ability for only albino shoot were used in symmetric fusion. On the other hand, the protoplasts of I. ensata and I. germanica protoplasts which were inactivated by iodoacetamide (IOA) treatment were used in asymmetric fusion. Five-six months after cell fusion, green plants were obtained in the symmetric and asymmetric fusion. In the random amplified polymorphic DNA (RAPD) analysis, the green plants had bands specific to both parental species. Therefore, these plants were somatic hybrids between I. ensata and I. germanica. This revised version was published online in July 2006 with corrections to the Cover Date.     

烟草原生质体再生植株的影响因子

原生质体培养和植株再生技术是植物生物工程的基础，植物原生质体的来源、酶解的条件、培养基的组成、培养方法等对原生质体的培养有很大的影响。本文着重对影响烟草原生质体培养再生植株的相关因素进行了综合论述，并讨论了原生质体培养技术在植物育种上的应用潜力。     

棉属G染色体组野生棉种的体细胞胚胎发生与植株再生研究

通过培养基激素配比、碳源种类等培养条件的筛选，对2个G染色体组棉种（纳尔逊氏棉，Gossypium nelsonii Fryx.；澳洲棉，Gossypium australe F. Muell.）进行体细胞培养并获得了体细胞胚，其中纳尔逊氏棉获得了再生植株，澳洲棉获得了大量的胚状体。与D染色体组棉种克劳茨基棉（Gossypium klotzschianum Anderss）相比，G染色体组棉种再生时间长，且体细胞胚畸形严重、萌发困难，但通过培养条件的调控可以得到大量胚状体和少量再生植株。激素组合0.1 mg L^-1 KT+0.1 mg L^-1 2, 4-D诱导的愈伤组织较松软，分化潜力高；胚性愈伤组织的增殖使用0.2 mg L^-1 KT+0.5 mg L^-1 IBA的激素组合；组合0.25 mg L^-1 IBA＋0.3 mg L^-1 KT有利于体细胞胚胎发生，而含激素组合MSB5+0.15 mg L^-1 KT+0.5 mg L^-1 NAA的培养基适合体细胞胚胎的萌发和植株再生。此外，愈伤组织诱导宜用葡萄糖作为碳源，而在胚性愈伤组织的增殖及保存和胚状体的萌发过程中用麦芽糖作碳源的效果更佳。     

Plant regeneration in sweet potato (Ipomoea batatas L., Convolvulaceae)

The application of new techniques for improvement of sweet potato crops, particularly including the exploitation of somaclonal variation, gene transfer by genetic transformation and somatic hybridization, requires the control of plant regeneration from tissue cultures. Shoots can easily be regenerated from explants of stems, petioles, leaves and roots, while callus cultures do not produce any shoots. The potential of somatic embryogenesis and plant regeneration via embryogenesis was evaluated for 10 cultivars of sweet potato. Protocols for plant regeneration from cultured protoplasts have also been developed. Since mesophyll was resistant to enzyme digestion, fragments of stems and petioles, callus and cell suspensions were used as source of protoplasts of sweet potato. Series of transfers of protoplast-derived calluses, particularly those which had been obtained from in vitro plants, to media containing a high level of zeatin resulted in successful formation of shoots in only two sweet potato cultivars. In addition, the embryogenic potential was irreversibly lost through protoplast culture, since protoplasts isolated from embryogenic cell suspensions developed into non-embryogenic callus. Consequently, an alternative protocol is being successfully developed to improve plant regeneration from cultured protoplasts of sweet potato, involving first root formation from which shoots can then be regenerated. Preliminary evaluation in field conditions in Gabon revealed that plants regenerated from cultured protoplasts exhibited a great genetic variability in their growth and tuber formation in particular. This revised version was published online in July 2006 with corrections to the Cover Date.     

Research in Tissue Culture of Diploid Cotton Species

Diploid cotton species provide parental (ancestral) species for tetraploid cultivars and are the key germplasm resources for cotton genetic improvement. In addition, diploid cotton species supply excellent materials for mutant library construction and functional genomics. Diploid cotton species originated from Asia- Africa, America, Oceania regions with rich genetic diversity. The tissue culture research mainly involves regeneration of plant organs and cell culture, protoplast culture, somatic culture and fusion. The regenerated plants have been obtained in Gossypium klotzschianum, G. gossypioides, G. arboreum, G. davidsonii, G. raimondii, G. stocksii, G. nelsonii, G. bickii, etc. Among them, regenerated plants in shoot tip culture were only obtained in G. arboreum, G. bickii. Regenerated plantlets in protoplast culture were obtained in G. klotzschianum, G. davidsonii. Interspecific hybrids between G. hirsutum and diploid cotton of G. klotzschianum, G. stocksii and G. bickii were obtained, respectively. The tissue culture systems of diploid cotton were different from each other, and the strategy is similar to that in woody plants. This paper summarized the progresses in related researches and discussed the future research.     

禾谷类作物原生质体培养研究进展

植物原生质体在品种改良、基因遗传转化、远缘杂交的应用研究以及生物学的基础理论研究中有着重要的研究价值。为了给禾谷类作物原生质体培养体系的建立提供理论依据,推动禾谷类作物品种遗传改良工作的开展,从原生质体制备、培养、再生、应用四个方面对禾谷类作物原生质体的培养研究进行了详细的阐述,分析了禾谷类作物原生质体制备和培养的主要影响因素,归纳了禾谷类作物原生质体再生的3种途径,对其在遗传改良和理论研究方面的应用进行了总结,并提出了禾谷类作物原生质体培养、遗传转化、细胞融合三个方面存在的问题和今后努力的方向。     

茶树叶片和胚根原生质体的分离及PEG诱导融合

植物原生质体是细胞培养和体细胞融合等细胞水平研究及植物遗传育种的重要材料。本研究用福鼎大白茶茶树的幼嫩叶片及胚根, 分析了原生质体分离过程中的材料、酶解液组成及酶解时间、纯化方法等影响因子, 建立了最佳原生质体分离体系, 为茶树体细胞杂交等细胞水平的研究提供了高效获取大量高活力原生质体的方法。结果表明, 23°C恒温黑暗或遮光培养的茶树实生苗的5周叶龄以内的幼嫩叶片是茶树原生质体分离的最佳材料, 其次是茶树种子萌发后的幼嫩胚根; 而以茶园健康生长的5周叶龄以内的幼嫩叶片为材料时, 只能获得混有大量细胞碎片的少量具有活力的原生质体。以茶树幼嫩叶片为分离材料的酶解液组成为1.5%纤维素酶+0.1%离析酶+0.5%果胶酶+0.4 mol L^-1甘露醇+20 mmol L^-1 MES; 以茶树幼嫩胚根为分离材料的酶解液组成为1.5%纤维素酶+0.3%离析酶+0.5%果胶酶+0.4 mol L^-1甘露醇+20 mmol L^-1 MES。分离茶树幼嫩叶和幼嫩胚根原生质体时, 宜采用低速(分别为55 r min^-1和50 r min^-1)恒温(23°C)摇床振荡酶解培养, 时间分别为7 h和8 h; 最适宜采用15×g的转速, 离心4 min可纯化获得高产量和活力的原生质体。用40% PEG-6000诱导20 min后可使茶树原生质体融合, 融合率达10%。     

陆地棉原生质体培养与植株再生技术研究

 分离培养陆地棉品种ZDM-3（Gossypium hirsutum L cv ZDM-3）胚性细胞悬浮系的原生质体,通过体细胞胚胎发生成功获得再生植株。试验结果表明,植物生长调节剂组合和碳源对原生质体培养具有重要的影响,添加2,4-D + KT + 1.5%（W/V）葡萄糖+ 1.5%（W/V）麦芽糖的KM8P培养基对于陆地棉原生质体培养的效果较好。激素组合0.46 μmol·L^-1 KT + 0.45 μmol·L^-1  2,4-D诱导的愈伤组织较松软,分化潜力高;胚性愈伤组织的增殖使用0.93 μmol·L^-1 KT + 2.46 μmol·L^-1 IBA的激素组合;1.2 μmol·L^-1 IBA + 1.39 μmol·L^-1 KT有利于体细胞胚胎发生,而MSB-5 + 0.67 μmol·L^-1 KT+2.69 μmol·L^-1 NAA的培养基适合体细胞胚胎萌发和植株再生。此外,愈伤组织诱导宜用葡萄糖作为碳源,而胚性愈伤组织增殖、保存和胚状体的萌发过程宜使用麦芽糖作碳源。     

植物原生质体再生细胞壁研究进展

原生质体作为单细胞且具有全能性,为研究植物细胞壁生物合成提供了独特的视角和模型.原生质体细胞壁再生过程复杂,涉及诸多细胞信号转导通路,应用普通的研究方法难以全面解析整个再生壁过程的分子网络.为充分了解植物原生质体再生壁内在特性,推动植物原生质体再生壁的分子机理进一步完善,笔者从原生质体再生细胞壁培养、细胞壁再生的细胞生...     

Plant regeneration from protoplasts of Iris germanica L.

Summary Protoplasts were isolated enzymatically from suspension cultures derived from embryogenic calli induced by leaf base culture of Iris germanica. In protoplast culture, the effects of glucose concentration, different sugars and combinations of 2,4-D and KIN on protoplast division and colony formation were examined. N6 medium supplemented with 0.1–1 mg/l 2,4-D, 1 mg/l KIN, 200mg/l casein hydrolysate, 250 mg/l proline, 0.2 M glucose and 20 g/l agarose was suitable for protoplast division and colony formation. When colonies formed were transferred onto hormone-free MS medium, many plantlets were regenerated through somatic embryogenesis. Thus, we could establish a plant regeneration system from protoplasts of I. germanica.Contribution from the Laboratory of Plant Breeding, Faculty of Agriculture, Miyazaki University, Japan, No. 95.