Somatic embryogenesis from styles of lemon (Citrus limon)

Lemon [Citrus limon (L.) Burm.] styles were treated with different growth regulators for induction of somatic embryos. Styles and stigmas were dissected from flowers and cultured on a Murashige and Skoog (MS) basal medium supplemented with 4.52 M 2,4-dichlorophenoxyacetic acid and 13.3 M 6-benzyladenine. Callus was induced from the style base 2 weeks after the treatment initiation, and embryos appeared 2 months later.Abbreviations BA 6-benzyladenine - 2,4-d 2,4-dichlorophenoxyacetic acid - MS Murashige & Skoog (1962) - NAA -naphthaleneacetic acid     

Continuous micropropagation of juvenile larch from different species via adventitious bud formation

A continuous propagation of juvenile larch in vitro was based on adventitious bud formation and different cytokinin combinations were tested concerning their effectiveness to induce elongation of adventitious buds. Zeatin (1.5 mg dm-3) combined with kinetin (0.15 mg dm-3) was found to be the best. Development and elongation of buds was achieved on a modified LP-medium. Using this system it was possible to propagate different larchspecies (Larix decidua, L. gmelinii and L. sukaczewii f. multiramosus) continuously. Shoots were successfully rooted and transferred to the soil. This revised version was published online in July 2006 with corrections to the Cover Date.     

Plant regeneration from embryo-derived tissue cultures of soybeans

Summary Routine regeneration of fertile plants from a tissue culture of soybean has been achieved. Serially propagated embryogenic cultures were initiated from immature embryos of many genotypes. Organized tissues developed only on the cotyledons of embryos. Genotypic variation in the frequency of initiation of embryogenic tissue was noted. However, embryogenic tissue cultures were generated from all genotypes tested. Embryogenic tissue was serially increased and underwent morphogenesis. Whole fertile plants were recovered. Cultures have been maintained for two years without loss of morphogenic competency. Editor's Statement This procedure for initiating embryogenic tissue cultures from commerical cultivars of soybean and the subsequent development of fertile plants establishes a framework for studying the processes of embryogenesis and embryogenyin vitro as well as providing a system for tissue culture propagation and in vitro modification of soybean. Robert B. Horsch     

Shoot regeneration from stem and leaf explants of Dendranthema grandiflora Tzvelev (syn. Chrysanthemum morifolium Ramat.)

Adventitious shoots were regenerated from leaf and stem explants of eleven chrysanthemum cultivars. The optimum medium for both explant types contained Murashige & Skoog basal medium supplemented with 5 M 6-benzylaminopurine and 5 M -napthaleneacetic acid. Generally, stem explants were superior to leaf explants. There were large cultivar differences in shoot regeneration frequency with three cultivars failing to respond over a wide range of hormone combinations. Shoots on stem explants appeared mainly to originate from cortical cells which rapidly divided and ruptured the epidermis. Regenerated shoots could be easily rooted, transferred to glasshouse conditions, and grown to flowering. All regenerated plants had the same morphological characteristics compared to plants derived from nodes.Supported in part by the National Biotechnology Program Research Grants Scheme in collaboration with Calgene Pacific P/L, Victoria, Australia.     

扶芳藤组织培养再生体系的建立

扶芳藤茎段比叶盘易产生不定芽;4种基因型中,圆瓣扶芳藤的不定芽再生频率远远高于其他3种;MS 6-BA 2.0mg·L-1是圆瓣扶芳藤不定芽诱导的最适培养基;低温处理10 d可显著提高不定芽的再生频率;暗培养对扶芳藤再生影响不大;扶芳藤诱导不定芽的最适pH范围在5.8～6.2之间;最适琼脂浓度为0.6%.     

4种红景天植物的组织培养研究

以大花红景天、云南红景天、长鞭红景天和库页红景天的茎和叶为外植体进行组织培养,结果表明：大花红景天以茎为外植体诱导芽效果最好,其它3种红景天以叶为外植体诱导芽效果最好。云南红景天和长鞭红景天适合的芽诱导激素组合是0．1mg／L NAA和2．5mg／L 6-BA的组合,在该激素水平下两种红景天的出芽频率分别达到71％和84％;大花红景天和库叶红景天适合的芽诱导激素组合是0．5mg／L NAA和2．5mg／L 6-BA的组合,在该激素水平下两种红景天的出芽频率均达到80％。长鞭红景天和库叶红景天在添加IBA的培养基上容易生根形成完整植株,生根率分别达到87％和73％;经过炼苗后,长鞭红景天再生苗能够成功移栽,成活率达66％。     

甜椒的离体培养及建立再生体系研究

以荷兰进口甜椒的子叶和下胚轴为外植体,接种到附加不同植物生长调节剂的培养基上,筛选出MS+6-BA 4.0 mg/L+NAA 0.1 mg/L为子叶和下胚轴最佳不定芽分化培养基;MS+IBA 0.2 mg/L和MS+IBA 0.5 mg/L为最佳不定根分化培养基。     

西兰花再生体系的建立与优化

目的:以西兰花无菌苗为材料,对影响西兰花再生的各种素进行研究,建立并优化西兰花的再生体系.方法:用组织培养法,对西兰花的外植体进行诱导生芽、生根.结果:不同品种的再生率差别较大,珠绿和青秀两个品种的再生率达91%以上,而鼎丰一号仅为80.3%;培养6 d 的下胚轴分化能力最强;当培养基中添加3 mg/L 6-BA 和0.2 mg/L IAA 时,下胚轴的分化率最高,玻璃化程度也降到最低;当培养基中添加0.2 mg/L IBA 或0.2 mg/L NAA时,可成功诱导不定芽生根.结论:建立并优化了西兰花再生体系,为西兰花遗传转化体系的建立奠定了基础.     

Somatic embryogenesis in clonal neem, <Emphasis Type="Italic">Azadirachta indica</Emphasis> A. Juss. and analysis for <Emphasis Type="Italic">in vitro</Emphasis> azadirachtin production

Summary Efficient and highly reproducible induction of somatic embryogenesis was obtained in four out of seven selected clones of neem, Azadirachta indica A. Juss. This was achieved either directly from root and nodal explants or indirectly from callus cultures initiated from leaf explants excised from 1-yr-old axenic plants. Direct induction of somatic embryogenesis was achieved both from nodal and root segments within 8 wk of culture on MS1 medium without growth regulators. However, the addition of 2.3–4.5 μM thidiazuron and 0.5 μM 2,4-dichlorophenoxyacetic acid into the medium were necessary to induce somatic embryogenesis via callus phase from leaf explants. Repetitive embryogenesis was observed within 3–4 wk following transfer of somatic embryos to a plant growth regulator-free medium. When somatic embryos of nodal and root segments were left on the induction medium without subculturing, approximately 15% of the somatic embryos developed into whole plantlets after passing through a series of developmental stages. Plantlets thus produced were hardy, lush green, and acclimatized casily under greenhouse conditions. However, somatic embryos derived from leaf explants showed low conversion rates (<5%). HPLC analysis revealed no detectable levels of azadirachtin in somatic embryos.     

In vitro shoot tip multiplication of cowpea Vigna unguiculata (L.) walp

Summary Shoot multiplication was induced in cowpea, cv. Georgia-21, from shoot tip explants. Shoot tips, 5 mm long, were isolated from in vitro-grown seedlings and cultured on MS medium containing N⁶-benzyladenine (BA) at 1, 2.5, or 5 mg/liter (4.4, 11.1, or 22.2 μM) or 6-furfurylaminopurine (kinetin) at 1, 2.5, or 5 mg/liter (4.6, 11.6, or 23.2 μM) combined with 2,4-dichlorophenoxyacetic acid (2,4-D) at 0.01, 0.1, or 0.5 mg/liter (0.05, 0.5, or 2.3 μM) or naphthaleneacetic acid (NAA) at 0.01, 0.1, or 0.5 mg/liter (0.05, 0.5, or 2.7 μM). Cultures were maintained at a 12-h photoperiod (40 μmol·m⁻²·s⁻¹) and 23 ± 2° C. Treatments with BA induced greater shoot proliferation than those with kinetin. The highest number of shoots was produced on 5 mg (22.2 μM) BA per liter in combination with NAA or 2,4-D at 0.01 mg/liter (0.05 μM). Callus proliferated from the basal ends of shoot pieces in all treatments. The cultures also formed roots in the presence of kinetin, but not on BA-containing medium. To produce whole plants, the shoots were separated and rooted on 0.1 mg (0.5 μM) NAA per liter. Resulting plants grew normally under greenhouse conditions. Shoot tips provide an excellent explant source for cowpea micropropagation and can be used for callus induction.     

不同红掌品种的叶片、叶柄和茎段愈伤组织的诱导及植株再生

红掌幼嫩叶片表面灭菌的最佳方法是：0．01％KMnO4 10min,0．05％链霉素、0．05％制菌霉素和0．05％头孢唑林钠各灭菌20min,然后用75％乙醇30s和HgCl2 2min,污染率为零。Anthurrium adraeanum “Rosetta“和A.adraeanum “Oilcloth-flower”愈伤组织诱导率和芽分化率显著高于A.adraeanum “Atlanta”.A.adraeanum “Fantasis”,A.adraeanum“Afrikanerin”。不同品种茎段、叶柄、叶片的愈伤组织诱导率和芽分化率有明显差异,大多数品种是幼茎段＞叶柄＞叶片。幼苗移栽在草煤或血竭渣基质中,成活率可达97％。     

盐生植物海马齿离体再生

建立盐生植物海马齿（Sesuvium portulacastrum）的离体再生体系,为其生物技术改良奠定基础。以海马齿叶片、茎和腋芽为外植体, 在不同激素配比的培养基上进行愈伤组织诱导、继代培养以及不定芽的分化和生根培养。结果表明： 最适愈伤组织诱导的外植体为叶片, 其次为幼嫩的茎段和腋芽。以叶片为外植体, 愈伤组织诱导率最高的培养基为MS＋2.0mg·L^–12, 4-D ＋ 0.5 mg·L^–16-BA ＋ 3%sucrose; 芽分化最适培养基为MS ＋ 1.0 mg·L^–1 2, 4-D ＋ 0.2 mg·L^–1 6-BA ＋ 3% sucrose;生根最适培养基为MS ＋ 3%sucrose ＋ 0.1%AC。炼苗移栽后, 成活率可达80%。     

Embryogenic response of Mexican alfalfa (Medicago sativa) varieties

The in vitro embryogenic response of nine varieties of alfalfa (Medicago sativa L.) grown in México (five Mexican varieties: Puebla 76, Inia 76, Bajío 76, Sintético I and Sintético II and four foreign or introduced varieties: Moapa 69, San Joaquín II, Hairy Peruvian and Valenciana) were tested. We screened 25 genotypes from each variety in four tissue culture protocols. All the varieties, except San Joaquín II, gave a positive response in one or more of the protocols tested. The response in each variety was low; this was also observed in a wider screening performed with the varieties Moapa 69, Hairy Peruvian, Sintético I and Sintético II. Two plants from Moapa 69 were regenerated and appeared normal.     

Adventitous bud induction in vitro from juvenile leaves of Cupressus arizonica Green

Juvenile leaves of Cupressus arizonica Green (3–5 mm in length) from eight week old seedlings, were cultured on liquid medium supplemented with isopentenyladenine (2 mgl^-1). Buds formed from the explants after three weeks of culture, but further growth occurred only after transfer to half-strength medium without plant growth regulators. Histological analysis at different times of culture, showed an early mitotic activity within transfusion tissue, followed by dedifferentiation of epidermal and mesophyll parenchyma cells at the basal zone of the leaves. The differentiation of vascular nodules always preceded bud formation. The difficulty of conifers to root and grow beyond plantlet stage is discussed.     

仙客来的离体无性试管繁殖研究

利用引种仙客来的叶片、叶柄、球茎等为外植体进行组织培养与再生植株 ,筛选最适诱导和继代增殖培养基为MS +BA 2 (mg/l,下同 ) +KT 0 .2 +2 ,4 -D 0 .5。其中叶片的诱导率和繁殖系数较高 ,叶柄次之 ;生根较难 ;球茎因常内带菌而造成培养困难     

Induction of morphogenically competent callus and suspension cell cultures from leaf explants and mature seeds ofDigitaria sanguinalis

Summary Digitaria sanguinalis (crabgrass) has recently been introduced as a high quality forage crop. We report here a tissue culture system showing a high level of regeneration developed to aid in a breeding program. Two morphologically distinct types of callus, compact opaque and friable translucent, were induced from leaf blade explants and mature seeds when cultured on MS medium containing 0.9 μM 2,4-dichlorophenoxyacetic acid. Proline (25 mM) inhibited induction of callus but was required for continued maintenance. Plants were readily regenerated from the compact opaque callus. Selectively subcultured friable translucent callus continued to produce colony sectors of the morphogenically competent compact opaque callus when transferred to regeneration medium. Suspension cell cultures derived from callus or directly from leaf blade explants also produced regenerable callus.     

芳香堆心菊离体再生体系的建立

芳香堆心菊(Helenium aromaticum)全株具芳香气味, 且头状花序仅含管状花, 是研究菊科植物花香和花型的良好材料, 但目前尚缺乏对其转基因技术体系的研究。为建立高效的芳香堆心菊离体再生体系, 以叶片、茎段和下胚轴为外植体, 进行25组不同激素及不同浓度配比的不定芽诱导研究。结果表明, 以芳香堆心菊叶片为外植体, 培养基为MS+ 0.2 mg·L^-1 NAA+1 mg·L^-1 6-BA+0.2 mg·L^-1 TDZ, 培养20天后愈伤组织诱导率高达100%, 丛生芽的诱导率为62.10%; 将不定芽接种于1/2MS培养基中进行生根培养, 16天即可生根, 且生根率为63.33%; 生根后继续培养14天现蕾, 开花率达93.33%。此外, 研究表明芳香堆心菊的再生受外植体来源、激素种类和浓度的影响。2,4-D不利于芳香堆心菊不定芽的诱导, 适宜浓度的6-BA和TDZ组合能有效促进芳香堆心菊不定芽的形成。研究初步建立了芳香堆心菊组织培养条件下的离体再生体系, 为建立其遗传转化体系奠定了坚实的基础。研究结果还可用于后续有关菊科植物花香和花型的研究。     

三叶半夏悬浮培养下的体细胞胚胎发生及植株再生

用三叶半夏幼嫩叶片诱导产生的胚性愈伤组织建立了胚性细胞悬浮系,研究了悬浮培养下体细胞胚胎的发生及植株的再生。结果表明,胚性悬浮细胞在附加1．0mg／L2,4-D、0．2 mg／LBA和300mg／L LH的MS液体培养基中产生大量的球形胚,转入液体分化培养基(MS 0．1 mg／LNAA 0．2 mg／L BA 300 mg／L LH)中进一步发育成心形胚、鱼雷形胚和子叶形胚。收集成熟胚转移到MS固体分化培养基上培养获得了再生植株。另外还观察到某些成熟胚上产生了许多次生胚。     

In vitro micropropagation and plant establishment of muscadine grape cultivars (Vitis rotundifolia)

Shoot apical meristems were used to establish regenerative axillary bud cultures of 9 muscadine grape cultivars. Meristems taken from 10 cm long shoots had less contamination (3%) and a higher survival rate (94%) than those from shorter or longer shoots. Of media tested, MS, 1/2 MS, and C₂D resulted in equivalent shoot proliferation rates, whereas, WPM produced stunted shoots. When pooling results for 3 cultivars, 5, 10 and 20 M BA and 5 M TDZ produced the highest average number of shoots per cultured apex (3.4–3.8). However, shoots produced with TDZ were stunted and did not root well. For rooting of shoots directly in potting mix, a rooting powder pretreatment significantly increased the number of roots per shoot but did not affect percent rooting or root length. For rooting in vitro, 1 M NAA significantly increased all parameters measured. Although more shoots rooted in vitro than in vivo (77% vs. 46%), the latter was judged preferable since acclimatized plants were produced in less time and a major culture step was eliminated. Significant differences among cultivars were noted for measured responses in all experiments.Abbreviations BA benzyladenine - Kin kinetin - MS Murashige & Skoog (medium) - NAA naphthaleneacetic acid - TDZ thidiazuron - WPM woody plant medium