番茄离体培养过程中器官发生的细胞组织学观察

对番茄下胚轴、子叶、茎段、叶片、叶柄不同类型外植体离体培养中有关细胞启动、分裂、分化以及器官发生作了细胞组织学观察。研究结果表明番茄不同类型外植体在同样的培养条件下,愈伤组织生长表现出明显差异,其中下胚轴、子叶诱导产生愈伤组织时,细胞启动最早,生长最快,其分裂方式基本为无丝分裂,未见有丝分裂,因此我们认为以不定芽方式获得转基因植株时,植株的所有性状变化,是否纯属目的基因所为,应该反复考察,不能忽视不定芽产生过程中的种种变化;下胚轴诱导愈伤组织形成时,细胞不规则的无丝分裂少于子叶,故下胚轴离体培养得到的正常芽的比例高于子叶的;番茄离体培养中不定芽通常发生在愈伤组织的周边区,也可起源于维管组织结节周围的形成层状细胞。不定根则由茎中柱鞘处发生。     

番茄离体培养过程中器官发生的细胞组织观察

对番茄下胚轴、子叶、茎段、叶片、叶柄不同类型外植体离体培养中有关细胞启动、分裂、分化以及器官发生作了细胞组织学观察。研究结果表明：番茄不同类型外植体在同样的培养条件下,愈伤组织生长表现出明显差异,其中下胚轴、子叶诱导产生愈伤组织时,细胞启动最早,生长最快,其分裂方式基本为无丝分裂,未见有丝分裂,因此我们认为以不定芽方式获得转基因植株时,植株的所有性状变化,是否纯属目的基因所为,应该反复考察,不能忽视不定芽产生过程中的种种变化;下胚轴诱导愈伤组织形成时,细胞不规则的无丝分裂少于子叶,故下胚轴离体培养得到的正常芽的比例高于子叶的;蕃茴离体培养中不定芽通常发生在愈伤组织的周边区,也可起源于维管组织结节周围的形成层状细胞。不定根则由茎中柱鞘处发生。     

枫香下胚轴的离体培养和植株再生

1　植物名称　枫香 (Ｌｉｑｕｉｄａｍｂａｒｆｏｒｍｏｓａｎａ)。2　材料类别　无菌种子萌发后的幼苗下胚轴。3　培养条件　 ( 1 )无菌种子萌发培养基 :ＭＳ ;( 2 )不定芽诱导培养基 :1 /2ＭＳ 6 ＢＡ0 .5、1 .0、2 .0ｍｇ·Ｌ- 1 (单位下同 ) ＮＡＡ 0 .1 ;( 3)不定芽增殖培养基 :1 /2ＭＳ 6 ＢＡ 1 .0 ＮＡＡ 0 .1 ＧＡ 30 .5 ;( 4 )生根培养基 :ＷＰＭ ＩＢＡ 2 .0。所有培养基蔗糖均为 3% ,琼脂 0 .65 % ,ｐＨ 5 .8,1 2 1℃高温、高压灭菌 1 6ｍｉｎ。培养温度 2 4～ 2 6℃ ,光照 1 6ｈ·ｄ- 1 ,光照度 2 0 0 0ｌｘ左右。4　…     

芥蓝下胚轴离体培养及高频率植株的再生

三个芥蓝品种下胚轴离体植株再生的条件的研究结果表明：下胚轴切口处可直接诱导出芽,诱导“早花尖叶芥蓝”、“中花尖叶芥蓝”和“迟花尖叶芥蓝”直接出芽的最佳激素组合分别为2．0mgL-1BA,0．3mgL-1NAA＋2．0mgL-1BA,0．5mgL-1NAA＋2．0mgL-1BA,其相应的芽发生频率分别为84．6％,86．7％,93．3％。诱导芽发生的最适蔗糖浓度是1％。培养基中加入4．0mgL-1AgNO3和500mgL-1MES可显著提高芽再生频率。再生芽在MS附加0．1mgL-1NAA的培养基上诱导生根形成完整植株。离体再生苗与种子萌发实生苗田间生长外形差别不大,但长势稍慢。     

暴马丁香下胚轴的离体培养和植株再生

1　植物名称　暴马丁香 (Syringareticulatavar.mandshurica)。2　材料类别　成熟蒴果种子的下胚轴。3　培养条件　 ( 1 )芽诱导培养基 :MS + 6 BA 5mg·L- 1 (单位下同 ) +IBA 0 .0 5～ 0 .5 ;( 2 )增殖培养基 :MS + 6 BA 5 +IBA 0 .1 ;( 3)生根培养基 :1 / 2MS +IBA 1。以上各培养基加 2 %蔗糖、0 .6%琼脂 ,高压灭菌前pH 5 .8。培养温度 2 5～ 2 8℃ ,光照度 1 5 0 0～ 2 0 0 0lx ,光照时间 1 6h·d- 1 。4　生长与分化情况4.1　种子的预处理　暴马丁香种子于秋季采收、晒干 ,4℃下贮藏两个月后 ,用洗涤剂洗净 ,纱布捆扎 ,流水冲…     

枣叶片离体培养再生植株

PlantletRegenerationfromLeavesCulturesofZizyphusiuiubaCHENZong－Li,YANZhi－Lian,QILong（Denyt17mllofmp,You’onl／nll＊,ndy,Yan’as716000）1植物名称枣（凯W…。Wwi）。2材料类别俗名“狗头枣”的无菌试管苗的叶片。3培养条件（l）叶片愈伤组织诱导及继代培养基：MS＋6－BA0．3mg／L（单位下同）＋2,4D20;（2）芽分化培养基：MS＋6－BAI．0＋IBA0．2＋D一泛酸钙1．0十活性发0．5％;（3）芽生长培养基：1／2MS＋6－BA0．2＋IAA0．04＋D一泛酸钙1．0;（4）芽增殖培养基：1／2MS＋6－BA0．4＋IAA0．0…     

山苍子的离体培养和植株再生

1　植物名称　山苍子 (Ｌｉｔｓｅａｃｕｂｅｂａ) ,又名木姜子。2　材料类别　成年植株带芽茎段。3　培养条件　所使用的培养基为 :( 1 )ＭＳ 6 ＢＡ3.0ｍｇ·Ｌ- 1 (单位下同 ) ＩＢＡ 0 .5 ;( 2 )ＭＳ 6 ＢＡ2 .0 ＩＢＡ 0 .5 ;( 3)ＭＳ 6 ＢＡ 1 .0 ＫＴ 1 .0 Ｎ     

山葵的离体培养和植株再生

CInvitroCultureandPlangtletRegenerationofEutremawasabiWANGGuang-Dong,LIShi-Jun(DepartmentofHorticulture,NanjingAgriculturalUniversity,Nanjing210095)1植物名称山葵（Eutremawasabi）,又名山嵛菜。2材料类别带侧芽的根状茎。3培养条件（1）诱导培养基：1／2MS＋6－BA0．2mp·L－1（单位下同）＋NAA0．05;（2）增殖培养基：MS＋6-BA0．2＋KT0．2＋NAA0．05;（3）生根培养基：1／2MS＋NAA0．05。培养基（1）附加琼脂粉5．0g·L－1,其余附加6.0g·L－1。（1）和（2）培养基加蔗糖30g·L－1,（3）加20g·L－…     

厚皮甜瓜的离体培养植株再生

1　植物名称　厚皮甜瓜 (Ｃｕｃｕｍｉｓｍｅｌｏｓｓｐ .Ｐａｎｇ)主栽品种“皇后”。2　材料类别　种子。3　培养条件　诱导胚性愈伤组织培养基 :( 1 )ＭＳ大量及微量元素 (下同 ) 6 ＢＡ 1 .5ｍｇ·Ｌ- 1 (单位下同 ) ;( 2 )ＭＳ Ｂ5 有机 6 ＢＡ 1 ;( 3)ＭＳ ＮＡＡ1 6 ＢＡ 1 ;( 4 )ＭＳ ＮＡＡ 1 6 ＢＡ 0 .5 ;( 5 )ＭＳ ＮＡＡ 1 6 ＢＡ 0 .1 ;( 6)ＭＳ ＩＡＡ 0 .1 6 ＢＡ 1。芽分化培养基 :( 7)ＭＳ 6 ＢＡ 0 .2 ;( 8)ＭＳ 6 ＢＡ0 .5 ;( 9)ＭＳ 6 ＢＡ 1 .0 ;( 1 0 )改良ＭＳ 6 ＢＡ 0 .2 ;( 1 1 )改良…     

Plant regeneration from cultured hypocotyl explants of diploid and tetraploid Solanum khasianum clarke

Plant regeneration through shoot bud formation was induced in cultured hypocotyl explants of diploid and tetraploid varieties of Solanum khasianum. Optimum regeneration of multiple shoot buds accompanied by rooting occurred on Murashige and Skoog's medium supplemented with IAA (1 mg/1) and kinetin or benzyladenine (1 mg/1). Plantlets obtained in vitro were successfully established in soil.Part of a thesis entitled Morphogenetic studies in tissue and organ cultures of certain Solanaceous plants: Capsicum, Lycopersicon and Solanum submitted by A.L. Gunay and approved for Master's degree in Botany by research of the University of Bombay.     

Organogenic regeneration of soybean from hypocotyl explants

Summary Thirteen soybean genotypes representing maturity groups IV−VI were compared for organogenic responses on three media cultured under two lighting conditions with hypocotyl sections excised from 7-d-old seedlings. All soybean lines responsed by producing adventitious shoots on the acropetal end of the hypocotyl explants, confirming genotype-independence of shoot initiation. Media containing 6-benzyladenine (BA; 5.0–10 μM) induced the greatest numbers of shoots. Histological studies confirmed the adventitious nature of arising shoots by indicative formation of meristematic zones and shoot primordia from parenchymatous tissues of central pith and plumular trace regions of the hypocotyl. Incompletely excised cotyledonary buds also contributed to shoot initiation. Degrees of responses were media-dependent and varied with regard to genotype. Centennial, Epps, and Lyon gave the greatest individual responses. Between cultivars (across all treatments), the regeneration potential (percentage of explants producing meristem-like structures or shoot primordia) 4 wk after initiation ranged from 47 to 75%. Four wk later, regenerative ability (number of shoots produced per responding explant) and regeneration efficiency (number of shoots produced per explant plated) yielded 1.4–7.1 and 1.0–5.0 shoots, respectively. The optimized protocol included initiation on a medium containing 5.0 μM BA for 4 wk, then transfer onto a shoot elongation medium (0.36 μM BA) for 4 wk. For 11 genotypes tested, 66–100% of excised shoots produced roots after 4 wk on media containing 12.5–29.2 μM indole-3-butyric acid. Of 109 regenerants transplanted to soil, 94% survived and no sterility has been observed on those mature enough to flower.     

Plant regeneration from sweet pepper (Capsicum annuum L.) hypocotyl explants

Morphogenetic potential of hypocotyl and cotyledon explants of the three Polish Capsicum annuum L. cultivars (Kujawianka, Passat and Zorza) was studied to develop a reliable plant regeneration protocol. Out of 8 and 15 combinations of growth regulators used in the first and second series respectively, the best medium contained BAP (5 mg·l⁻¹) and IAA (1 mg·l⁻¹). Media containing thidiazuron (TDZ) and IAA proved to be worse than those with BAP and IAA. Additionally, it was indicated that hypocotyl explants placed upside-down developed more adventious buds. ‘Passat’ was the most responsive variety (approximately 40 % of both types of explants produced buds). In the second part of experiment the aim was to stimulate shoot induction in the conditions most adapted to Agrobacterium transformation. ‘Bryza’ replaced cv ‘Kujawianka’ and proved to be better than ‘Passat’, previously distinguished as a highly responsive cultivar. The experiments confirmed a significant effect of the hypocotyl explant length and induction period on shoot regeneration. Finally, the optimum concentration of carbenicillin and kanamycin was determined.     

Efficient regeneration from hypocotyl explants in three cotton cultivars

A high frequency in vitro shoot bud differentiation and multiple shoot production protocol from hypocotyl segments of 8 to 10-d-old seedlings of cotton has been developed. Murashige and Skoog (MS) basal medium with Nitsch and Nitsch vitamins was found to be optimal in shoot regeneration. A combination of 2 mg dm⁻³ thidiazuron and 0.05 mg dm⁻³ naphthaleneacetic acid was the most effective for shoot regeneration (76 %) and an average of 10.6 shoots per responding explant. Combination of the cytokinins benzylaminopurine and kinetin induced better regeneration response than their individual treatments. Supplementation of the culture medium with ethylene inhibitor silver nitrate and activated charcoal showed beneficial effects. Optimal rooting was obtained on half-strength MS medium supplemented with 1 mg dm⁻³ indolebutyric acid and activated charcoal. Scanning electron micrographs of in vitro cultured explants revealed that shoot primordia were formed de novo.     

Endophytic fungi in leaves of Lycopersicon esculentum Mill

Endophytic fungi were isolated from healthy tomato leaves from field-grown plants. Several surface-sterilization techniques were assayed for removing and killing epiphytes on tomato leaves. Surface-sterilization with an undiluted commercial solution of sodium hypochlorite was employed. To determine the spectrum of endophytes, samples were collected in February 1998 and 1999 from tomato plants (cultivar Tommy) cultivated in the experimental fields of Facultad de Ciencias Agrarias y Forestales, Universidad Nacional de La Plata, Buenos Aires, Argentina. In general, different spectra of species were isolated in 1998 and 1999.Alternaria alternata was the fungus most frequently isolated from tomato leaves in 1999, but it was the second most common species in 1998. In contrast, Colletotrichum gloeosporioides was the fungus most frequently isolated in 1998, but it was not found in 1999. Species of other genera, such as Cladosporium and Penicillium, were isolated in both years.     

In vitro plantlet regeneration from cotyledon, hypocotyl and root explants of hybrid seed geranium

In vitro plantlet regeneration systems for the seed geranium (Pelargonium x hortorum Bailey) using cotyledon, hypocotyl and root explants were optimized by studying the influence of seedling age, growth regulators and excision orientation on organogenesis. Indole-3-acetic acid combined with zeatin yielded the highest rate of shoot production on cotyledon explants (0.2–2 shoots per explant). More shoots were produced on explants cut from the most basal region of cotyledons from 2 to 4-day-old seedlings than from older seedlings or more distal cut sites. Hypocotyl explants produced the highest number of shoots, up to 40 shoots per explant, on indole-3-acetic acid (2.8–5.6 mM) + zeatin (4.6 mM) or thidiazuron (4.5 mM). Maximum shoot formation (0.3–1.4 shoots per explant) on root explants occurred when they were cultured on medium containing zeatin. Regenerated shoots rooted best on a basal medium containing no growth regulators. There were substantial differences among cultivars in shoot formation from each of the explant systems.Abbreviations BA 6-benzylaminopurine - 2,4-d 2,4-dichlorophenoxyacetic acid - IAA indole-3-acetic acid - NAA naphthaleneacetic acid - TDZ thidiazuron     

Pluripotency of Arabidopsis xylem pericycle underlies shoot regeneration from root and hypocotyl explants grown in vitro

We have established a detailed framework for the process of shoot regeneration from Arabidopsis root and hypocotyl explants grown in vitro . Using transgenic plant lines in which the GUS or GFP genes were fused to promoters of developmental genes ( WUS , CLV1 , CLV3 , STM , CUC1 , PLT1 , RCH1 , QC25 ), or to promoters of genes encoding indicators of the auxin response ( DR5 ) or transport ( PIN1 ), cytokinin (CK) response ( ARR5 ) or synthesis ( IPT5 ), or mitotic activity ( CYCB1 ), we showed that regenerated shoots originated directly or indirectly from the pericycle cells adjacent to xylem poles. In addition, shoot regeneration appeared to be partly similar to the formation of lateral root meristems (LRMs). During pre-culture on a 2, 4-dichlorophenoxyacetic acid (2, 4-D)-rich callus-inducing medium (CIM), xylem pericycle reactivation established outgrowths that were not true calli but had many characteristics of LRMs. Transfer to a CK-rich shoot-inducing medium (SIM) resulted in early LRM-like primordia changing to shoot meristems. Direct origin of shoots from the xylem pericycle occurred upon direct culture on CK-containing media without prior growth on CIM. Thus, it appeared that the xylem pericycle is more pluripotent than previously thought. This pluripotency was accompanied by the ability of pericycle derivatives to retain diploidy, even after several rounds of cell division. In contrast, the phloem pericycle did not display such developmental plasticity, and responded to CKs with only periclinal divisions. Such observations reinforce the view that the pericycle is an 'extended meristem' that comprises two types of cell populations. They also suggest that the founder cells for LRM initiation are not initially fully specified for this developmental pathway.     

Nitrosomethylurea induced streptomycin resistance in Lycopersicon esculentum Mill

High frequency of streptomycin resistant variants of Lycopersicon esculentum were isolated on selective shoot regeneration medium supplemented with IAA (0.5 mg/L), zeatin (1.5 mg/L) and streptomycin sulphate (500 mg/L). Nonmutagenized (controls) and NMU treated cotyledons were placed on shoot regeneration medium supplemented with antibiotic streptomycin. Resistant shoots appeared at a high frequency in mutagenized cotyledons, whereas in controls morphogenesis was suppressed, accompanied by bleaching. Shoot regeneration occurred from the nodular tissues developed at the cut ends of cotyledons. Resistant shoots developed into complete plantlets on rooting medium containing selective concentration of antibiotic. Stability of streptomycin resistance was confirmed by leaf assay and reciprocal crosses between streptomycin-resistant and sensitive plants.