伊贝母体细胞无性系的建立及其胚状体的发生

本文报道了伊贝母体细胞无性系的建立及其胚状体的发生。已继代培养三年零六个月共30多代的鳞芽愈伤组织,目前仍有分化能力。通过愈伤组织形态细胞学的观察,发现伊贝母体细胞无性系形成小鳞茎的途径有二:一是由特化了的愈伤组织表皮细胞。经多次分裂发育成不定芽而形成小鳞茎;二是由愈伤组织表层或内层特化了的胚性细胞,经多次分裂发育成胚状体而形成小鳞茎。不定芽和胚状体的形态发生是有区别的。     

油茶子叶体细胞胚形成的细胞学观察

以油茶子叶块为外植体,在附加2.0mg·L-1 2,4-D和1.0mg·L-1KT的MS培养基上进行培养,观察诱导产生的体细胞胚性愈伤组织细胞学结构.结果表明:油茶体细胞胚可直接起源于表皮或近表皮的单细胞原胚或者多细胞团.其中,单细胞原胚先分裂形成二细胞原胚,二细胞原胚再进一步分裂后聚集形成多细胞团,最终经过球形胚、梨形胚、心形胚发育形成一个完整的体细胞胚.     

水稻愈伤组织内部胚性细胞的形成及发育

发育成胚状体的水稻愈伤组织表面胚性细胞, 在继代过程中被非胚性细胞旺盛的分裂所包围, 渐渐形成内部胚性细胞。其形态结构与表面胚性细胞相同, 但周围缺乏表面细胞下面的一层至几层含丰富的淀粉粒的细胞。内部胚性细胞形成团后先在以处形成突起, 产生根冠原, 逐渐形成根而突出愈伤组织。内部胚性细胞可向四周同时但不同步形成根冠原。未见芽或胚状体从内部胚性细胞产生。推测胚性愈伤组织失支胚胎发生能力及分化能力可能部分地与胚性细胞部分裂或分裂不旺盛、而非胚性细胞分裂旺盛从而使胚性细胞被包围、稀释有关。     

茴香组织培养中体细胞胚胎发生的组织细胞学研究

将茴香幼茎或叶柄的愈伤组织转入附加６－ＢＡ和低浓度２,４－Ｄ的ＭＳ培养基以后,愈伤组织逐步由松软状转变成为颗粒状的胚性愈伤组织。胚状体起源于胚性愈伤组织中的单个细胞或胚性细胞团。在含ＮＡＡ和６－ＢＡ的培养基中,胚状体发育成熟,并再生小植株。茴香的胚状体主要以单细胞内起源方式发生。首先由胚状体单个原始细胞分裂形成２－细胞原胚,２－细胞原胚以三种方式进行分裂：１。Ｔ－形分裂;２．直线形分裂;３．田字形     

福录考体细胞胚胎发生的观察

福录考（ＰｈｌｏｘｄｒｕｍｍｏｎｄｉｉＨｏｏｋ）体细胞胚胎发生的方式有两种：一是由叶外植体具有一定功能的特化细胞脱化为胚性细胞,如叶表皮细胞,维管束鞘及韬皮薄壁细胞均可发生脱分化进行分裂,形成胚性细胞或分细胞团;二是叶外植体先脱分化形成愈伤组织。     

黄山栾树体细胞胚的发生和组织学观察

黄山栾树无菌苗的节间和叶柄离体培养后,其体细胞胚发生的结果表明：节间愈伤组织可诱导产生体细胞胚,而叶柄愈伤组织则生根：节间愈伤组织诱导培养基为MS＋3．0mg．L～2,4．D＋0．5～3．0mg．L-1NAA;节间胚性愈伤组织诱导培养基为MS＋2．0nag．L-2,4-D;胚性愈伤组织转移到无植物生长调节剂的MS培养基上可发育成正常植株。组织学观察表明,体细胞胚在胚性愈伤组织中有的发生于愈伤组织表层细胞,有的发生在愈伤组织内部。黄山栾树体细胞胚的形成经历球形胚、心形胚、鱼雷胚和子叶胚几个阶段,这与合子胚的发育途径相似。     

水稻愈伤组织内部胚性细胞的形成及发育

发育成胚状体的水稻愈伤组织表面胚性细胞,在继代过程中被非胚性细所包围,渐渐形成内部胚性细胞。共形态结构与表面胚性细胞相同,但周围缺乏表面细胞下面的一层至几层含丰富的淀粉粒的细胞,内部胚性细胞形成团后行然以外形成突起,产生根冠原,逐渐形成根而突出愈伤组织,内部胚性细胞可向四周同时但不同步形成根冠原,未见芽或胚状体从内部胚性细胞产生,推测胚性愈伤组织失支胚胎发生能力及分化能力可能部分与胚性细胞部分裂或分裂不旺盛、,而非胚性细胞分旺盛从而使胚性细胞被包围,。稀释有关。     

香雪兰的直接与间接体细胞胚胎发生

香雪兰的体细胞胚胎发生可通过两种途径进行,即直接发生与间接发生。在直接发生方式中,体细胞胚直接来源于尚未完全分化的外植体表皮细胞;体细胞胚与母体组织以一种类似胚柄的结构相联系。间接发生方式中,体细胞胚的形成要经过一个愈伤组织阶段。以是否能形成体细胞胚分类,可将愈伤组织分为胚性和非胚性愈伤组织。以间接方式形成的体细胞胚是由胚性愈伤组织中的一种决定细胞发育来的。这种体细胞胚不具有类似胚柄的结构,而与母体组织共同形成一个复合体。体细胞胚具有自己独立的维管束系统,在脱离母体组织后能够独立发育成株。     

高粱体胚发生的组织细胞学观察

以Sb33高粱非胚性、胚性愈伤组织和体胚为材料,用传统石蜡切片法对各组织材料进行组织化学染色,对高粱胚性与非胚性愈伤组织以及体胚进行组织细胞学观察。结果表明：高粱非胚性愈伤组织无淀粉粒积累,高粱胚性愈伤组织淀粉粒积累较多,而与胚性愈伤组织相比,高粱体胚淀粉粒积累更多,这说明淀粉粒的积累与高粱体细胞的胚胎发生密切相关。此外,高粱可通过鱼雷胚基部产生球形胚的方式实现体胚的增殖,高粱离体再生途径以体细胞胚发生为主,并同时存在少量器官发生途径。在高粱体细胞胚胎发生中,外起源和内起源同时存在。本研究为高粱体细胞胚胎发生提供细胞学理论基础。     

黄连体细胞胚胎发生的研究

黄连(Coptis chinensis)叶片外植体在 MS 2,4-D 1 ppm 培养基上很容易产生愈伤组织。愈伤组织在转入分化培养基 MS 6-BA 0.5ppm NAA 1ppm 培养基上以后,能产生大量胚状体。胚状体可经过球形、心形、鱼雷形及子叶期等诸阶段发育成小植株。对胚状体用4%的藻酸钠和2%的氯化钙进行人工种皮包埋后,在无菌条件下,胚状体转变成苗。愈伤组织在分化培养基上经几次继代后,整个愈伤组织可转变为胚性愈伤组织并形成一个个胚性细胞团。胚状体可从其表面或愈伤组织内的任一细胞团产生。这一研究结果为获得大量分散的单个胚状体及人工种子的研制提供了良好的实验系统。     

Plant regeneration through somatic embryogenesis from suspension cultures of a minor millet,Paspalum scrobiculatum

Compact, friable and embryogenic calli were initiated from immature inflorescences and young leaf bases of one week old seedlings of Paspalum scrobiculatum cultured on MS medium supplemented with 2,4-D. A stable, embryogenic suspension culture was initiated from these calli and maintained in a liquid version of the same MS medium. Embryogenic calli and somatic embryos were obtained by plating suspension culture cells onto semi-solid medium containing 2,4-D. Complete, normal plantlets developed on 2,4-D free medium at a high frequency from somatic embryos. NAA and BAP in the medium promoted plant development.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - NAA 1-naphthaleneacetic acid - BAP 6-benzylaminopurine - ABA Abscisic acid - MS Murashige and Skoog (1962) - CM Coconut milk     

High Frequency of Plant Production via Somatic Embryogenesis from Callus or Cell Suspension Cultures inEleutherococcus senticosus

Hypocotyl segments ofEleutherococcus senticosuscultured on Murashigeand Skoog's (MS) medium with 4.5 µM2,4-D produced somaticembryos directly from the surface of explants without interveningcallus formation. When these somatic embryos were subculturedto the same MS medium with 4.5 µM2,4-D, friable embryogeniccalli were formed mainly from radicle tips of somatic embryos,but at a low frequency (5%). Selected embryogenic calli weremaintained on MS agar or liquid medium with 4.5 µM2,4-D.To induce somatic embryo development, embryogenic calli andcell clumps were transferred to MS medium lacking 2,4-D. Thefrequency of somatic embryo formation differed between culturetypes with 1570 embryos formed per Petri dish from callus cultureand 5514 embryos formed per flask from cell suspension cultures.Somatic embryos formed on agar medium had larger cotyledonsthan those of embryos formed in liquid medium. GA₃treatmentwas necessary to induce germination from somatic embryos. Therate of plant conversion was 97% in somatic embryos from callusculture and 76% in embryos from liquid culture. Regeneratedplantlets were successfully acclimatized in the glasshouse.Copyright1999 Annals of Botany Company Eleutherococcus senticosus, micro propagation, somatic embryogenesis.     

Plant Regeneration from Protoplasts of Peucedanum Terebinthaceum (Fisch.) Fisch. ex Turcz

Calll with many embryogenic cell colonies were produced from segments of seedlling of Peucedanum terebinthaceum (Fisch.) Fisch. ex Turcz. which were cultured on the 1/2MS agar medium (with half quantity of macronutrients) containing 1 mg/l 2,4-D. Cell suspension culture with high percentage of embryogenic cell colonies was established from the calli shaking in liquid medium. The cell suspension culture was used for protoplast preparation. Protoplasts were obtained with the enzyme mixture containing 1.5% Onozuka R-10, 0.3% Macerozyme R-10, 0.5% Snailase, 5 mmol/l CeCl2, 1 mmol/l KH2PO4, 0.6 mol/l mannital at pH 5.8 and 25℃. Cultured in a modified MS liquid medium containing 1 mg/l 2,4-D+ 0.5 mg/l zeatin, the protoplasts emered division after four days, and formed cell colonies of 0.5–1mm after about forty days. When transfered to 1/2 MS liquid medium supplemented with zeatin (0.5 mg/l), the cell colonies differentiated in to embryoids, then developed into plantlets with many green leaves and roots on the 1/2 MS agar medium devoid of phytohormones.     

新疆雪莲体细胞胚胎发生

通过体细胞胚胎发生途径实现了新疆雪莲（Saussurea involucrata Kar．et Kir．）的植株再生。选用新疆雪莲子叶为外植体,接种于MS＋0．5mg·L^-12,4-D＋0．05—1mg·L^-1BA的固体培养基上,进行愈伤组织的诱导。从第1次继代培养的愈伤组织中挑选出黄绿色、颗粒状、质地致密的腔陛愈伤组织,转移到含0．05—0．1mg·L^-1 2,4-D的MS液体培养基中进行悬浮培养,20天后可分化产生大量球形胚。继代过程中相继加入PEG和GA3,可以促进体细胞胚的分化和生长。体细胞胚在含有5mg·L^-1 GA3的MS固体培养基上,可发育成完整的植株。     

悬浮培养的谷子细胞的胚胎发生和植株再生(简报)

由谷子的胚性愈伤组织在附加2mg/l2,4-D和5%椰乳的UM液体培养基中建立了细胞悬浮培养,降低培养基中2,4-D的浓度,利于胚状体的形成。当液体培养中的细胞转移到MS琼脂培养基上后,通过改变激素的组成及浓度,可以促进胚性细胞团的增殖,进而再生出大量完整植株。这种通过形成胚状体而再生植株的能力,巳在该悬浮培养系中保持一年多,从由幼穗培养建立胚性愈伤组织开始,此细胞系的旺盛的再生能力至今巳保持了近三年。     

Relationship between auxin-induced cell proliferation and somatic embryogenesis in culture of carrot cotyledons

We elucidated the relationship between cell proliferation and somatic embryogenesis in the culture of carrot cotyledons. Fresh weights of the cotyledon expiants were determined every five days while being cultured on a medium containing 2,4-D. Callus production increased exponentially from Day 20 to Day 25, showing a two-fold rate of proliferation. To examine the embryogenic potential of the callus, we pre-cultured cotyledon explants on an MS medium with 2,4-D, then transferred them to an MS basal medium at five-day intervals. Somatic embryos formed most frequently when the cotyledons were pre-cultured for 20 days on an MS medium that contained 5 μ2,4-D. The frequency of somatic embryo formation was 81%, while that of normal embryos with two cotyledons was 51% among those formed on a hormone-free medium. We used FACScan analysis to relate the embryogenic potential of the callus to the S phase in the cell cycle of cultured cells. The S phase was high after 25 days of culture on the medium with 5 μM 2,4-D. In contrast, the frequency of normal embryogenesis was higher at Day 20 of the pre-culture period. Culturing embryogenic calli on a medium with 5 μM 2,4-D was most favorable for producing somatic embryos with two cotyledons. We verified that active somatic embryogenesis was apparently related to cell division activity; somatic embryos induced from actively dividing cells were apt to accompany cotyledonary abnormality.     

Direct somatic embryogenesis from protoplasts of Foeniculum vulgare

Protoplasts prepared from an embryogenic cell suspension culture of fennel gave rise to somatic embryoids directly through unequal cell divisions of enlarged, ellipsoidal cells, when embedded in hormone-free LS agarose medium. On the other hand, protoplasts embedded in LS agarose medium containing 2,4-D and kinetin proliferated through unpolarized cell divisions to form calli, which gave somatic embryoids on the surface upon transfer onto the same medium. In either case, somatic embryoids germinated to develop into normal plantlets when cultured on hormone-free LS agar medium under illumination.     

Somatic embryogenesis and plant regeneration from cell suspension culture of niger (Guizotia abyssinica Cass.)

Somatic embryogenesis was achieved from cell suspension cultures of niger (Guizotia abyssinica Cass.). Initially, friable embryogenic calluses were induced from cotyledonary leaves of niger on Murashige and Skoog (MS) agar medium containing 5 μM 2,4-Dichlorophenoxyacetic acid (2,4-D) and 0.5 μM kinetin (KIN). Cell suspension cultures were established by using embryogenic calluses in MS liquid medium containing 5 μM 2,4-D and 0.5 μM KIN. Initiation of somatic embryogenesis and development up to globular stage from embryogenic cell clumps occurred in the liquid medium itself. Thereafter embryogenic cell aggregates were transferred to MS agar medium supplemented with 3 μM KIN for embryo differentiation, whereas maturation of somatic embryos occurred in MS agar medium containing 10 μM abscisic acid.     

Plant regeneration via somatic embryogenesis from solid and suspension cultures of Vitis vinifera L. cv. ‘Manicure Finger’

Vitis vinifera L. cv. ‘Manicure Finger’ is one of the major table grape varieties in China. To provide a strong foundation for genetic transformation with potential for crop improvement, we undertook plant regeneration via somatic embryogenesis. Anthers and gynoecia were harvested from immature flowers and used as explants to induce embryogenic calli. Explants cultured in MS1 medium (based on Murashige and Skoog basal salts), supplemented with 4.5-μM 2,4-dichlorophenoxyacetic acid (2,4-D) and 4.4-μM 6-benzylaminopurine (6-BA) showed the highest rates of embryogenic callus induction (3.7%?±?1.3% for anthers and 4.8%?±?2.5% for gynoecia). After several months, somatic embryos were produced from embryogenic calli cultured in plant growth regulator-free MS2 medium (with reduced sucrose). Somatic embryos (SE) at the cotyledonary stage were isolated and cultured on three different media (MS2, MS3, or B) for conversion into plantlets, the efficiency of which ranged from 63.9%?±?4.8% to 83.9%?±?8.4%. After 1 mo of in vitro culture, 80% of plants with at least six leaves were successfully transplanted into soil. SE was repeatedly induced from previously induced somatic embryos for up to 1.5 yr. Using embryogenic calli as starting material, suspension cultures containing embryogenic cell aggregates were also established in liquid MS medium supplemented with 4.5-μM 2,4-D. The embryogenic cell aggregates continued to proliferate without differentiating for successive subculture cycles. After transfer to 2,4-D-free liquid medium for 4 wk, an average of 63.7%?±?9.0% mature SEs were produced per 20 mL of liquid medium. More than 40% of somatic embryos at cotyledonary stage, derived from the suspension cultures, successfully germinated into plants using solid medium.     

新疆雪莲体细胞胚胎发生

通过体细胞胚胎发生途径实现了新疆雪莲(Saussurea involucrata Kar.et Ki r.)的植株再生。选用新疆雪莲子叶为外植体, 接种于MS+0.5 mg.L-1 2,4-D+0.05-1 mg.L-1 BA的固体培养基上, 进行愈伤组织的诱导。从第1次继代培养的愈伤组织中挑选出黄绿色、颗粒状、质地致密的胚性愈伤组织, 转移到含0.05-0.1 mg.L-1 2,4-D 的MS液体培养基中进行悬浮培养,20天后可分化产生大量球形胚。继代过程中相继加入PEG和GA3 , 可以促进体细胞胚的分化和生长。体细胞胚在含有5 mg.L-1 GA3 的MS固体培养基上, 可发育成完整的植株。