几种匍匐翦股颖成熟胚愈伤组织诱导及植株再生

利用成熟种子作为外植体,分析了2,4-D对匍匐翦股颖胚性愈伤组织诱导与植株再生体系的影响,并对体细胞胚的发生过程进行了观察.实验结果表明,在2.0 mg/L 2,4-D 0.1 mg/L 6-BA时胚性愈伤组织的诱导频率最高.随着2,4-D浓度的增加,愈伤组织的诱导和分化能力明显下降.在再生过程中采用1.0 mg/L的6-BA达到了比较好的效果,愈伤组织的再生频率大部分在90%以上.同时发现适当提高肌醇浓度可以使苗长得较为粗壮.在实验中发现匍匐翦股颖的体细胞胚发生过程为球形胚、心形胚、鱼雷胚和子叶胚.     

荷兰芹胚性愈伤组织诱导及胚状体发生与内源IAA和ABA关系的初步研究

分析了荷兰芹胚性愈伤组织发生的条件, 对Co²⁺作用下胚状体形成与培养物内源IAA和ABA 的关系做了研究。结果表明, 荷兰芹下胚轴胚性发生能力随其相对位置而异, 自下而上逐渐提高。提高2, 4-D 浓度有利于胚性愈伤组织的诱导, 水解酪蛋白对胚性能力的表达没有显著影响。在胚状体发生过程中, 添加Co²⁺显著降低培养物内源IAA 和ABA 水平, 并提高胚状体诱导率。其中对照组IAA 有一个峰值, ABA 有两个峰值;实验组ABA 变化趋势与对照组相似, 而IAA 则始终没有峰值出现。Co²⁺对IAA 和ABA 的抑制机制可能有所不同。     

玉米幼穗离体培养体细胞胚高频发生的研究

以玉米（Ｚｅａ ｍａｙｓ Ｌ．）幼穗为材料,ＭＳ主基本培养基,诱导体细胞且状体。筛选出了胚状体高频发生的品种;同时实验结果表明：玉米遗传型与胚状体形成有一定的相关性;诱导期２,４－Ｄ浓度对且状体产生十分重要,ＫＴ和６－ＢＡ影响且状体的发生。组织切片观察表明：多数胚状体起源于愈伤组织表层,少数产生于里层;愈伤组织产生且状体是不同步的。     

百合体细胞胚胎发生和植株再生

以切花百合(Lilium)品种‘黄天霸’(‘Manissa’)花器官为外植体诱导体细胞胚胎发生与植株再生。结果表明,不同花器官、不同激素配比对愈伤组织形成均具有显著影响。花丝为最佳外植体,激素对愈伤组织诱导的影响效应为NAA>6-BA>2,4-D,最适培养基为MS+1.0 mg.L-1NAA+0.2 mg.L-16-BA;激素诱导体细胞胚胎发生的影响效应为2,4-D>KT>6-BA,最佳培养基配方为MS+1.0 mg.L-12,4-D+0.2 mg.L-1KT+1.0 mg.L-16-BA;MS培养基添加IBA可促进体细胞胚萌发成苗,体细胞胚芽成苗的最佳培养基为MS+0.2 mg.L-16-BA+1.0 mg.L-1IBA。     

Plant regeneration through somatic embryogenesis in callus culture of green bamboo (Bambusa oldhamii Munro)

Summary Young inflorescence explants of green bamboo (Bambusa oldhamii Munro) in culture show a high capacity for plant regeneration through somatic embryogenesis. Embryogenic callus was initiated from explants maintained on Murashige and Skoog's medium supplemented with 3 mg/l 2,4-D, 2 mg/l kinetin and a high content (60 g/l) of sucrose. Prolonged culture in the embryoid induction medium or transferral of embryonic callus to auxin-free medium resulted in the continued development and eventual germination of embryoids and establishment of rooted plantlets that were successfully transferred to soil.     

Cytological and Embryological Studies on Haploid Plant Production from Cultured Unpollinated Ovaries of Nicotiana tabacum L.

We obtained mature haploid (n = 24) ovary plants from in vitro cultured unpollinated young ovaries. These ovaries were induced to form embryoids which then developed into plants. The results obtained are summarized as follows: 1. The origin of development of the ovary haploid plants has been followed by light microscopy. Embryological abservations revealed that there are two ways of plantlet production: (1) Ovary haploid plant was derived from the macrospore without an intervening callus phase. (2) Ovary haploid plant was derived directly from the egg cell of mature embryo sac. In addition, Callus derived haploid plant was also obtained from the base and the tip of a bud of the above mentioned haploid plantlet. In same medium embryoids was derived from callus. Finally, plantlet was developed. 2. The exogenous hormones are necessary for high induction frequency of embryoid from unpollinated isolated young ovary, but these are not definitely necessary for induction of embryonic callus to form embryoids which then developed into plant. 3. The induction frequency of embryoid from in vitro cultured ovary and embryonic callus significantly increased when the concentration of thiamine, pyridoxine, ascorbic acid, nicotinic acid, inositol and folic acid was raised.     

Evaluation of haemoglobin (erythrogen): for improved somatic embryogenesis and plant regeneration in cotton (Gossypium hirsutum L. cv. SVPR 2)

Somatic embryogenesis in cotton (Gossypium hirsutum L.) is accelerated when the plant regeneration medium is supplemented with haemoglobin (erythrogen). In cotton SVPR 2 lines, a higher frequency of embryoid formation was observed when the medium contained 400 mg/l haemoglobin. Fresh weight of the callus, rate of embryoid induction, number of embryoids formed and the percentage of plant regeneration from somatic embryos were increased. Among the two different cultivars tested, MCU 11 showed no response to the presence of haemoglobin when compared to SVPR 2, and embryogenic callus formation was completely absent in the former. Medium containing MS salts, 100 mg/l myo-inositol , 0.3 mg/l thiamine-HCL, 0.3 mg/l Picloram (PIC), 0.1 mg/l kinetin and 400 mg/l haemoglobin effected a better response with respect to embryogenic callus induction. After 8 weeks of culture, a high frequency of embryoid induction was observed on medium containing MS basal salts, 100 mg/l myo-inositol, 0.3 mg/l PIC , 0.1 mg/l isopentenyl adenine, 1.0 g/l NH₄NO₃ and 400 mg/l haemoglobin. Plant regeneration was observed in 75.8% of the mature somatic embryos, and whole plant regeneration was achieved within 6–7 months of culture. The regenerated plantlets were fertile and similar to in vivo-grown, seed-derived plants except that they were phenotypically smaller. A positive influence of haemoglobin was observed at concentrations up to 400 mg/l at all stages of somatic embryogenesis. The increase in the levels of antioxidant enzyme activities, for example superoxide dismutase and peroxidase, indicated the presence of excess oxygen uptake and the stressed condition of the plant tissues that arose from haemoglobin supplementation. This increased oxygen uptake and haemoglobin-mediated stress appeared to accelerate somatic embryogenesis in cotton.Abbreviations BAP Benzylaminopurine - 2,4-D 2,4-Dichlorophenoxyacetic acid - GA₃ Gibberellic acid - GR Glutathione reductase - 2iP Isopentenyl adenine - KT Kinetin - NAA Naphthaleneacetic acid - PFC Perfluorocarbon - PIC Picloram - PO Peroxidase - ROS Reactive oxygen species - SOD Superoxide dismutase - T.HCl Thiamine hydrochloride     

Cytology and Histology in Somatic Embryogenesis of Indica Rice

The somatic embryogenesis was established from mature dehulled seeds. The histological research showed that embryogenic calli were initiated first from absorbed cells of scutellum of mature seed. And then the embryoids derived from the surface of embryogenic callus. Having been the same structure like a zygotic embryo of rice, the embryoids possessed the major parts of scutellum, coleoptile and coleorhiza. In an embryoid, several developmental stages of pro-embryoid, including single embryogenic cells, two, four and multiple cell stage pro-embryeids and some abnormal embryoids were observed. It could be concluded from this experiment that the embryoid from somatic cell culture in Indica rice possessed an original form of a plant in structure like a zygotic did and derived from a single cell.     

盐肤木体细胞胚胎发生及植株再生

以盐肤木(Rhus chinensis Mill.)幼胚为外植体,研究不同植物生长调节剂组合对其愈伤组织诱导及体细胞胚胎发生的影响,以建立盐肤木体细胞胚胎发生及植株再生体系。结果表明,最适愈伤组织诱导培养基为MS+6-BA 0.2 mg/L+2,4-D 1.0 mg/L,诱导率为84.57%,诱导出的初代愈伤组织白色或淡黄色,质地疏松,表面光滑,为非胚性愈伤。初代愈伤组织转移到1/2 MS+6-BA 2 mg/L+NAA 0.5 mg/L培养基上培养1个月后,长出淡黄色质地紧密的胚性愈伤组织,诱导率高达100%,在此培养基上胚性愈伤组织增殖倍数为854.73%。所获得的胚性愈伤组织转接到1/2 MS+6-BA 2 mg/L+NAA 0.5 mg/L+蔗糖4%的培养基上培养1个月后可诱导体细胞胚胎发生,诱导率可达32.67%。诱导得到的体细胞胚胎经历球形胚、心形胚、鱼雷胚、子叶胚进一步分化发育成苗。无菌苗炼苗后栽种到泥炭土∶蛭石∶珍珠岩为2∶1∶1的生长基质上,能100%稳定成活。经过细胞学观察分析,体细胞胚的发育与合子胚相似。     

华山松胚性愈伤组织诱导与幼胚离体培养

探索华山松(Pinus armandii)体细胞胚胎发生技术对其实施规模化无性繁殖和开展遗传转化具有重要意义。本文以1/2LM为基本培养基, 通过激素调节等措施对华山松的胚性愈伤组织诱导和幼胚的离体培养技术进行了初步研究。研究结果: 胚性愈伤组织诱导率最高可达52.71%, 但愈伤组织继代培养后没有体细胞胚胎的分化; 首次从其子叶期的幼胚中直接诱导出具有根和茎的完整植株, 诱导率达92%以上。文章确认了采集的幼胚发育状态对胚性愈伤组织的诱导有重要影响, 并对诱导的培养条件等进行了探讨。     

华山松胚性愈伤组织诱导与幼胚离体培养

探索华山松（Pinus armandii）体细胞胚胎发生技术对其实施规模化无性繁殖和开展遗传转化具有重要意义。本文以1／2LM为基本培养基,通过激素调节等措施对华山松的胚性愈伤组织诱导和幼胚的离体培养技术进行了初步研究。研究结果：胚性愈伤组织诱导率最高可达52．71％,但愈伤组织继代培养后没有体细胞胚胎的分化;首次从其子叶期的幼胚中直接诱导出具有根和茎的完整植株,诱导率达92％以上。文章确认了采集的幼胚发育状态对胚性愈伤组织的诱导有重要影响,并对诱导的培养条件等进行了探讨。     

Morphogenesis in Long-term Maintained Immature Embryo-derived Callus of Wheat (Triticum aestivum L) - Histological Evidence for Both Somatic Embryogenesis and Organogenesis

Callus induced from immature embryos of wheat cv Kharchia 65 on Murashige and Skoog’s medium containing 2.5 mg l¹ 2,4-D was maintained In the regenerable state by subculturing every 5-6 weeks on medium supplemented with 2,4-D (2.5 mg l^-1) and proline (10 mg l^-1). Addition of proline helped maintain morphogenic competence for over four years. The regenerating callus was analysed histologically about one year after first induction. Both somatic embryogenesis and shoot organogenesis were seen in the same callus tissue that contained typical stages of somatic embryoid development and evidence for the de novo shoot bud formation.     

不同杂种优势类群玉米幼胚愈伤组织的诱导及植株再生特性的研究

以Reid、唐四平头和其它种质等3个杂种优势类群共19份玉米自交系为试验材料,以玉米幼胚作为外植体,研究了基因型、培养基和激素对玉米幼胚愈伤组织的诱导及植株再生的影响,结果表明供试材料均能进行愈伤组织的诱导,但是仅有12个自交系能再生植株。N6和改良N6培养基有助于提高愈伤组织的质量及其生长速度,2,4-D在愈伤组织的诱导中起着关键性作用。在诱导培养基中添加0.2mg／L的6-BA或KT会使胚性愈伤组织的诱导频率下降以及降低愈伤组织的质量。在胚状体诱导培养基中添加1mg／L的KT能促进绿苗的分化,但是浓度过高会使丛生苗分化过多。此外,通过对不同杂种优势类群自交系玉米幼胚培养特性的分析,发现在唐四平头类群的4个自交系中,黄早四的绿苗分化率仅为0.5％,其它3个自交系不能再生植株。但是,从Reid和其它种质类群的供试自交系中筛选出了胚性愈伤组织的诱导频率和绿苗分化率均较高的、适合于遗传转化的受体材料,如3189／4380、4380／陕综5、8103、先早17、18-599红、18-599白、501、178和冀53。     

暴马丁香成熟胚愈伤组织和体胚诱导及其生理状态解析

以暴马丁香成熟胚为外植体进行愈伤组织和体胚发生诱导,通过调节诱导培养基中植物生长调节剂的种类和浓度,分析其对愈伤组织诱导和体胚诱导的影响,同时对培养过程中的外植体进行形态发生观察和生理状态分析。结果表明:①暴马丁香成熟胚外植体培养30 d可见直接体胚发生、60 d可见子叶型体胚;②BA在愈伤组织诱导过程中起到了主导作用,在0.5 mg·mL^-1BA和5~6 mg·mL^-1NAA组合下,愈伤组织诱导率可达100%;在0.5 mg·mL^-1BA和5 mg·mL^-1NAA组合体胚诱导率可达8%;③多酚含量在愈伤组织形成初期急剧上升且在培养过程中保持较高水平,子叶型胚期PAL和POD活性升高、MDA和SOD活性略下降。     

细枝木麻黄离体培养过程中愈伤组织和再生芽的细胞组织学观察

为探讨细枝木麻黄(Casuarina cunninghamianaMiq.)愈伤组织分化过程的细胞组织学,对离体培养条件下的愈伤组织进行扫描电子显微镜和石蜡切片观察,分析愈伤组织的细胞分裂、分化以及芽再生的发生过程。结果表明,新鲜外植体培养于愈伤组织诱导培养基上,伤口处的薄壁细胞开始脱分化,培养1周后形成明显的愈伤组织;继续培养2周后,胚性愈伤组织形成,且表层细胞启动分化形成芽原基;培养4周,可肉眼观察到胚性芽原基,数量增多并逐渐分化形成不定芽;培养至第6周,生成不定芽,并大量增殖和分化。因此,细枝木麻黄是通过愈伤组织分化形成胚状体的途径进行植株再生的,为建立细枝木麻黄组织培养高效再生体系提供了理论依据。     

枇杷叶片胚性愈伤组织诱导与内源激素含量的关系

为揭示胚性愈伤组织发生过程中内源激素的变化规律,本研究以枇杷叶片为实验材料,通过诱导胚性愈伤组织获得体细胞胚,采用高效液相色谱法测定枇杷叶片及愈伤组织的赤霉素（GA₃）、生长素（IAA）、脱落酸（ABA）和玉米素（ZT）4种内源激素的含量,探讨胚性愈伤组织发育过程中4种内源激素的动态变化。结果显示,不同成熟度叶片内IAA/ZT比值对胚性愈伤组织的发生有正效应,而GA₃/IAA的比值具有负效应。胚性愈伤组织的发生需要较低含量的GA₃及高含量的IAA和ABA,IAA/ZT比值高有利于胚性愈伤组织形成,培养后期及时添加一定量的外源激素有利于胚性保持。本研究可为离体培养时选择外植体、添加外源激素及控制继代时间提供理论指导,并为快速获得枇杷胚性材料、开展基因转化研究奠定基础。     

Studies on Morphological Differentiation of Endosperm Plantlets of Chinese Gooseberry in Vitro

The present report deals with the process of embryoid induction and plantlet formation from cell-type endosperm of Actinidia chinensis var. chinensis and A. chinensis var. hispida. The callus was induced from endosperm on MS basic medium supplemented with zeatin 3 ppm, 2,4-D 0.5 ppm and CH 400 ppm and then transferred to differentiation medium of MS supplemented with zeatin 1 ppm and CH 400 ppm. After about half a month, the embryoid appeared from callus and then developed into plantlets. It could be seen from the histological figure 14—15, the embryoid of Chinese gooseberry is linear-shaped and consists of cells arranged in a long line in callus. When the cells regenerated at botk ends of the linear-shaped embryoid, the polarity of the embryoid is easily distinguished. The plantlets produced from embryoid appear rather stout at first and after some time, they changes gradually into normal plantlets.     

基于响应面设计茎段形成层培养猕猴桃幼苗的优化研究

为降低猕猴桃组培快繁中的污染率,提高其繁殖效率,该文以猕猴桃的幼嫩茎段为外植体,采用两步培养法进行茎段形成层的愈伤及成苗诱导研究,并利用响应面设计软件对NAA浓度、6-BA浓度、低渗处理时间进行了各条件的优化,同时通过组织切片确定愈伤的来源及幼苗的形成方式。结果表明:(1)培养过程中撕除茎段周皮能显著降低污染率,用200～400 mg·L^-1的PVP处理猕猴桃茎段可有效防止去皮茎段的褐化。(2)愈伤诱导的最佳条件为预培养28.3 h、NAA 4.45 mg·L^-1、6-BA 0.28 mg·L^-1,而幼苗形成的最佳条件为预培养26.4 h、NAA 4.84 mg·L^-1、6-BA 0.42 mg·L^-1。这表明形成层愈伤诱导需较长低渗处理时间和较高生长素,而成苗诱导则需较高生长素、激动素及较短的低渗处理时间。(3)组织切片观察结果表明猕猴桃愈伤组织源于形成层干细胞的分裂,且幼苗株源于胚状体的发育。综上结果表明,通过除去猕猴桃嫩茎周皮,外加抗氧化、低渗处理,可有效降低猕猴桃组培快繁中的污染率,提高繁殖系数和胚状体发生率,为猕猴桃种苗的规模化生产提供技术支撑。     

籼型杂交稻亲本成熟胚愈伤组织再生体系的建立

本文给出了6个籼型杂交稻亲本成熟胚愈伤组织再生体系建立优化措施。采用6个有重要育种价值的杂交籼稻亲本成熟胚盾片诱导愈伤组织作为分化再生的外植体,通过调节2,4-D浓度、培养基成分、接种方式、激素组合和愈伤组织分化途径,建立适合籼稻遗传转化的再生体系。结果表明,MB培养基是较为合适的愈伤组织诱导培养基类型,6个品种在MB培养基上的愈伤组织诱导率均在60％-80％之间。半粒米的接种方式能够明显提高愈伤组织诱导率,提高幅度达到28．2％。通过调节2,4-D浓度和激素组合,愈伤组织诱导率最高可达到97．9％,两步分化法和适当干燥处理能够提高愈伤组织的分化效率,6品种愈伤组织分化率均在50％-90％之间。初步建立了6个杂交籼稻亲本品种成熟胚愈伤组织的再生体系,为以后遗传转化奠定基础。     

外源激素对金钱松胚外植体愈伤组织的诱导及其器官发生的调节作用

金钱松胚外植体在培养过程中由于外源激素的种类和配比的不同而存在着几种发育途径：直接从胚外植体表面分化不定芽;先诱导愈伤组织,再从愈伤组织分化不定芽;还可由愈伤组织分化出胚状体。激素ＢＡ对外植体不定芽的诱导起着关键作用。激素２,４－Ｄ则诱导愈伤组织,ＢＡ与２,４－Ｄ配比恰当诱导的愈伤组织分化出体细胞胚状体。　ＬＰ’附加低浓度的ＢＡ或ＫＴ（＜０．５ｍｇ／Ｌ）促进不定芽茎的伸长;　ＬＰ’附加浓度的ＩＢＡ（＜０．５ｍｇ／Ｌ）诱导不定根的发生。愈伤组织在基本培养基浓度为　×ＬＰ’或１×ＬＰ’的分化培养基上不定芽诱导率相似。