湿地松体细胞胚胎发生和植株再生

以湿地松的未成熟合子胚为外植体,在附加８ｍｇ／Ｌ,２,４－Ｄ和４ｍｇ／Ｌ ＢＡ的ＬＰ培养基上诱导出胚性愈伤组织。在含１ｍｇ／Ｌ,２,４－Ｄ和０．５ｍｇ／Ｌ ＢＡ的培养基上保持并增殖。提高培养基的渗透压后,愈伤组织内大量的胚性胚柄细胞团和早期原胚。     

鱼腥草体细胞胚胎发生和植株再生

目的:利用鱼腥草的叶片和叶柄为材料,进行体细胞胚胎诱导及植株再生研究。方法:运用正交设计试验,考察在改良的MS固体培养基上添加不同种类、不同浓度的植物生长物质组合及其配比对鱼腥草愈伤组织诱导、体细胞胚胎发生及植株再生的影响。结果:鱼腥草无菌苗叶片在含有2,4-D 1.0 mg/L 6-BA 0.5 mg/L的改良MS培养基上能诱导出胚性愈伤组织;胚性愈伤组织在含有6-BA 1.0 mg/L的改良的MS培养基上诱导体细胞胚的发生;叶柄在含有6-BA 1.0 mg/L改良MS培养基上直接产生体细胞胚。体细胞胚在改良的MS NAA0.1 mg/L 6-BA 1.0 mg/L的培养基上能够快速繁殖,形成大量不定芽,在不加任何激素的MS培养基上就可以萌发出不定根,发育为成完整植株,在MS IBA 1.0 mg/L的固体培养基上能够形成大量的根。结论:建立了鱼腥草体细胞胚胎发生及植株再生的体系。     

一品红体细胞胚胎发生与植株再生

一品红不同部位愈伤组织诱导能力存在差异,嫩茎>幼花序>嫩叶。愈伤组织的长势主要受生长素的影响,细胞分裂素对愈伤组织生长有促进作用;但在含6-BA和NAA的培养基中诱导出的愈伤组织,其胚性明显强于单独用NAA诱导出的愈伤组织。液体悬浮培养是一品红体细胞胚胎高频发生的中间步骤。不同浓度BA对一品红体细胞胚的萌发率影响不大,萌发培养基中KNO₃含量加倍可提高萌发率。     

木薯体细胞胚胎发生及植株再生研究

对木薯体细胞胚胎发生的影响因素进行了优化研究。结果表明,基因型对木薯体细胞胚胎发生影响很大,在供试的六个品种中,“华南 8 号”的体细胞胚胎发生率和产胚量最高,分别为 65% 和19个;侧芽茎尖为最佳外植体,体细胞胚胎发生的最佳培养基为MS +0.5mg/L CuSO4 + 4 mg/L 2,4-D。同时,对木薯体细胞胚再生成完整植株的主要影响因素作了分析,建立了一个高效的植株再生体系。     

白Qian体细胞胚胎发生及其植株再生

白Ｑｉａｎ的成熟胚在４－６℃低温下保存１个月后,接种于改良ＬＰ＋２ｍｇ／Ｌ２,４－Ｄ＋１ｍｇ／Ｌ６－ＢＡ的培养基上,黑暗条件下培养１个月便可产生白色半透明的胚性愈伤组织。整体染色封片观察表明,胚性愈伤组织由很多很长的胚柄细胞及其顶端胚细胞团组成,这种愈伤组织培养物称为胚性胚柄团。     

水母雪莲体细胞胚胎发生及其植株再生

水母雪莲（Saussurea medusa Maxim.)茎和叶片的切段接种于MS＋2mg/L NAA 0.5mmg/L 6-BA的培养基上,20d后产生黄褐色的愈伤组织,经过几个月的继代培养,愈伤组织仍保持旺盛的增殖能力,但部分由黄褐色逐渐变为红色,将红色愈伤组织转到MS＋0．1mg/L NAA＋0．2mg/L 6-BA 5mg/L GA3的培养基上,30d后可分化出大量的体细胞胚,体细胞胚成熟后转到1／2MS＋0．2mg/L IAA 0.5%活性炭的培养基上,30d后可长出2－4cm的根,带根的小苗经锻炼后移栽到土壤中,成活率达76％,细胞组织学观察表明,发育成熟的体细胞胚具有胚根,胚轴和胚芽的完整结构,具有独立的维管系统。     

白杄体细胞胚胎发生及其植株再生

白杄(Picea meyeri Rehd.et Wils.)的成熟胚在4～6℃低温下保存1个月后,接种于改良LP 2mg/L 2,4-D 1mg/L 6-BA的培养基上,黑暗条件下培养1个月便可产生白色半透明的胚性愈伤组织。整体染色封片观察表明,胚性愈伤组织由很多很长的胚柄细胞及其顶端的胚细胞团组成,这种愈伤组织培养物称为胚性胚柄团。胚性胚柄团在MS 1mg/L 2,4-D 1mg/L KT的继代培养基上黑暗条件下可保持旺盛的增殖能力和分化潜力。当胚性胚柄团转到MS 5mg/L ABA 5mg/L AgNO_3的分化培养基上,1个月后可产生大量正常的体细胞胚。体细胞胚成熟以后转到含0.5％活性炭的无激素1/2MS基本培养基上约40d后可长出1.5～2.5cm的根,约60d后可长出真叶。光、ABA、蔗糖及AgNO_3浓度是影响体细胞胚发生的主要因素。     

玉米芽尖培养中的高频率体细胞胚胎发生与植株再生(简报)

通过诱导玉米芽尖产生胚性愈伤组织,建立起高频率植株再生的玉米芽尖培养实验体系。在MS+1.0mg·L-16-BA+0.2mg·L-12,4-D+500mg·L-1CH的培养基上诱导愈伤组织并继代培养1次后,将愈伤组织转移到Ms+0.5mg·L-16-BA+0.4mg·L-1IBA+500mg·L-1CH的分化培养基上,可形成大量的体细胞胚胎。组织学观察表明,体细胞胚胎主要发生在胚性愈伤组织表面与表层下部。不同基因型的芽尖形成愈伤组织和再生植株的能力不同。     

红肉小果型番木瓜品种'美中红'体胚的诱导

用红肉小果型番木瓜品种‘美中红’为外植体,探讨不同成熟度的幼胚、不同浓度2,4-D和培养条件对其体胚的诱导以及体胚形成过程的结果表明:以子叶和内外种皮都为白色且个体较大的番木瓜幼胚(90~120d)在含10mg·L-12,4-D的培养基中和黑暗条件下诱导愈伤组织的效果最佳,愈伤组织的诱导率随着2,4-D浓度的增加而增加。番木瓜愈伤组织最先发生于形态学上的胚根下端,体胚多发生于形态学上的胚芽上端。     

ACC oxidase from Carica papaya: Isolation and characterization

Most of the studies done on 1‐aminocyclopropane 1‐carboxylic acid (ACC) oxidase were done in vivo. It is only recently that in vitro studies have been carried out successfully on the enzyme. Here we report on in vitro studies of the enzyme that was isolated from Carica papaya . The enzyme had a K_m of 37 µ M and was inhibited by n ‐propyl gallate (0.240 m M ), sodium dithionite (0.022 m M ), sodium metabisulphite (0.021 m M ) and cobalt sulphate (0.100 m M ). The activity of the enzyme increased with ripening, the enzyme was somewhat labile and activity was lost after 4 days at 14°C; activity was prolonged when the crude homogenate was kept at −15°C. Isolation and purification were achieved with ammonium sulphate precipitation followed by gel‐filtration (Sephadex G 100‐120) and ion‐exchange chromatography (DEAE‐Sephadex). Gel electrophoresis of the purified enzyme gave a single band which corresponded to a molecular mass of 27.5 kDa. The amino acid content of the enzyme showed a relatively high percentage of valine (10.4%). Enzyme activity was enhanced when dithiothreitol (3 m M ) and bicarbonate ion (30 m M ) were added to the assay medium. The production of ethylene from Carica papaya did not require pretreatment of the fruit with ethylene.     

白杄体细胞胚胎发生及其植株再生条件的优化研究

为建立白木千体细胞胚胎发生及其植株再生的高频率实验系统 ,对聚乙二醇及干化处理的影响进行了系统研究。结果表明 ,在分化培养基中附加 5 0 g/L聚乙二醇可显著提高愈伤组织的分化频率和每块愈伤组织产生的体细胞胚个数 ,而干化处理又能使体细胞胚的萌发率大幅度上升     

番木瓜的离体繁殖

建立番木瓜离体繁殖体系.用0.1%HgCl2溶液对番木瓜的新生嫩茎进行消毒,适宜的消毒时间为12min,1mm茎尖的成苗率达到87.6%.随蔗糖浓度的提高,番木瓜试管苗的株高显著降低,增殖系数显著增加.在附加IBA0.3mg/L的1/2MS培养基上新梢的生根率达到89.3%,试管苗大规模移栽的成活率达90%以上.基因型显著地影响番木瓜离体增殖的效率.     

番木瓜核型和减数分裂研究

对番木瓜核型和花粉母细胞减数分裂行为的研究表明,番木瓜染色体数目为2n=18,由9对中部着丝粒染色体组成。核型公式为2n=2x=18m。花粉母细胞减数分裂正常,在终变期和中期Ⅰ观察到9个二价体,未观察到染色体结构变异和行为异常。     

Somatic embryogenesis from cell suspension cultures in Carica candamarcensis

Cell suspension cultures of Carica candamarcensis derived from hypocotyl calli were tested concerning their in vitro embryogenic capacity to improve asexual propagation rates in this species. Somatic embryos developed in culture from cells in suspension or from microcalli. Responses were affected by nutrient media and phytohormones used. Best results were obtained by growing the cells in suspension in Nitsch and Nitsch medium containing naphthaleneacetic acid and then plating them upon the same medium containing benzyladenine, or combinations of both hormones.     

Laticifer Ultrastructural and Immunocytochemical Studies of Papain in Carica papaya

Cotyledons of germinating papaya (Carica papaya L. ) seeds and exocarp of young fruits were used as materials for study. The ultrastructural changes occurring during differentiation of laticifer and the ultrastructural environment of papain synthesis were studied by means of TEM and immunocytochemistry. Electron microscopic observations showed that the differentiating laticiferous cells were rich in ribosomes and mitochondria. Endoplasmic reticulum (ER) was well developed and apparently active, forming secretory vesicles of various sizes. With further development, organelles were gradually degenerated and autophagy of cytoplasm within vacuole was evident. ER was dilated and split into fragments. Cell wall perforations occurred at several sites of adjacent laticifer elements. Towards maturity, laticifer was fully filled with vesicles containing electron-dense materials. Organelles disappeared thoroughly but plasmalemma remained. Sections were incubated with anti-chymopapain antibodies followed by goat-anti-rabbit IgG-gold. Labeled gold was found predominantly in ER and the associated vesicles of differentiating laticifer. Several controls were used to establish the specificity of the immunolaheling pattern. Investigations led to the conclusions that ER and polyribosomes were involved in papain synthesis. Papain was stored in the vesicles of ER origin temporally before reorganized into laticiferous vesicles with other components of latex.     

Somatic embryogenesis and plant regeneration from immature zygotic embryos of papaya (Carica papaya L.)

Summary Immature zygotic embryos from open-pollinated and selfed Carica papaya L. fruits, 90 to 114 days post-anthesis, produced 2 to 20 somatic embryos on apical domes, cotyledonary nodes, and radicle meristems after culture for three weeks on half-strength Murashige and Skoog (MS) medium supplemented with 0.1 to 25 mg l^–1 2,4-dichlorophenoxyacetic acid (2,4-D), 400 mg l^–1 glutamine, and 6% sucrose. After six weeks of culture, about 40 to 50% of the zygotic embryos had become embryogenic, and each embryogenic embryo yielded hundreds of somatic embryos within five months of culture on media supplemented with 2,4-D. Somatic embryos matured on half-strength MS medium, germinated on MS medium containing 5 mg l^–1 kinetin, and grew large enough for greenhouse culture on MS medium. Shoots were rooted in vermiculite and grown in the greenhouse.Journal Series no. 3449 of the Hawaii Institute of Tropical Agriculture and Human Resources     

A protocol for efficient transformation and regeneration of Carica papaya L.

Summary A reproducible and effective biolistic method for transforming papaya (Carica papaya L.) was developed with a transformation-regeneration system that targeted a thin layer of embryogenic tissue. The key factors in this protocol included: 1) spreading of young somatic embryo tissue that arose directly from excised immature zygotic embryos, followed by another spreading of the actively growing embryogenic tissue 3 d before biolistic transformation; 2) removal of kanamycin selection from all subsequent steps after kanamycin-resistant clusters were first isolated from induction media containing kanamycin; 3) transfer of embryos with finger-like extensions to maturation medium; and 4) transferring explants from germination to the root development medium only after the explants had elongating root initials, had at least two green true leaves, and were about 0.5 to 1.0 cm tall. A total of 83 transgenic papaya lines expressing the nontranslatable coat protein gene of papaya ringspot virus (PRSV) were obtained from somatic embryo clusters that originated from 63 immature zygotic embryos. The transformation efficiency was very high: 100% of the bombarded plates produced transgenic plants. This also represents an average of 55 transgenic lines per gram fresh weight, or 1.3 transgenic lines per embryo cluster that was spread. We validated this procedure in our laboratory by visiting researchers who did four independent projects to transform seven papaya cultivars with coat protein gene constructs of PRSV strains from four different countries. The method is described in detail and should be useful for the routine transformation and regeneration of papaya. Based in part on a presentation at the 1997 SIVB Congress on In Vitro Biology held in Washington, DC, June 14–18, 1997.