Phototropism and geotropism in hypocotyls of cress (Lepidium sativum L.)

Abstract. The phototropic and the geotropic sensitivities of cress hypocotyls differed in etiolated and in green seedlings. In etiolated seedlings, phototropism was poorly developed and geotropism seemed to be the dominant orientation mechanism. In green seedlings, although geotropic sensitivity was slightly greater, phototropic responses were markedly enhanced, such that photo-signals could over-ride, or even reverse, geocurvature. The 'de-etiolation' light pretreatments required to bring about these changes in the photo-and geo-systems were different. The slight enhancement of geocurvature could be achieved by short-term exposure to red light and was reversed by far-red irradiation. The marked stimulation of photocurvature required extensive irradiation pretreatments.     

A protocol for rapid micropropagation of endangered Isoplexis

Summary An efficient protocol for large-scale micropropagation of Isoplexis canariensis (L.) Loud., I. chalcantha Svent, and O'Shan., and I. isabelliana (Webb and Berth.) Masf. (Scrophulariaceae) is reported. Multiple shoots were obtained from shoot tips and nodal segments isolated from seedlings when cultured under varying conditions. Factors such as nutrient media, concentration of growth regulators, and type of induction medium (liquid or solid) strongly influenced shoot proliferation and development. Multiple well-developed shoots were obtained after induction on solid media containing Murashige and Skoog full-strength salts and low cytokinin concentrations. By changing the major salt formulation to half strength and employing liquid media the morphogenic parameters evaluated were affected negatively. Many shoots rooted spontaneously in hormone-free media and plants grew successfully in the greenhouse. Flowering was observed 6 mo. after ex vitro cultures were established.     

A protocol for Ulmus minor Mill. micropropagation and acclimatization

Here we report the establishment of a simple protocol for the micropropagation and acclimatization of U. minor. Branches with dormant buds were collected from mature elms and sprouted in a greenhouse. Tip and node segments were used as starting material for in vitro proliferation in a medium (designated here as DKW1) already used for the micropropagation of a clone of the English Elm (U. procera SR4). In the first assay, in which explants from nine different trees were used, 88.5% of the tip segments produced new axillary shoots thus proving to be the best explant type. Afterwards, material from four different trees (F4, F7, F13, F14), that had the highest sprouting rate in the greenhouse, was used to test for genotype influence. F14 proved to be the best genotype in culture and it was used for all the subsequent experiments. Shoots from F14 were used to assay in vitro rooting using five DKW based media. Rooting percentages were high for all media and varied between 80% and 100%. For acclimatization two approaches were assayed: the use of previously rooted in vitro plants and the direct acclimatization of shoots from cultures in DKW1. After 6 weeks, 86.4% of the in vitro rooted plants were successfully acclimatized and a slightly higher value, 88.6%, was attained by direct acclimatization of shoots with thick stems and hard leaves. These results proved that there is no need for a previous in vitro rooting step and that direct acclimatization can effectively reduce time and costs. Thus U. minor micropropagation and acclimatization can be divided into only two steps: proliferation of shoots in DKW1 and direct acclimatization of these shoots in a sterile soil mixture.     

In vitro micropropagation of the tropical fruit tree Syzygium cuminii L.

Multiple shoots were obtained from nodal and shoot tip segments of 10 to 15-day-old seedlings of Syzygium cuminii L. on Murashige & Skoog (MS) revised medium supplemented with 6-benzyladenine (BA) (0.23–8.90 M) singly or in combination with -naphthaleneacetic acid (NAA), indole-3-acetic acid (IAA) or indole-3-butyric acid (IBA). Excised shoots were placed for root induction on MS medium containing NAA and/or IBA and then transferred to MS basal medium to form complete plantlets. The regenerated plantlets have been acclimatized and successfully transferred into the soil.     

Culture of meristem tips and micropropagation of 12 commercial clones of poplar in vitro

Twelve commercial clones of poplar were cultured in vitro from meristem lips (0.3–0.5 mm diameter), shoot tips (4–6 mm long) and nodal segments (5–10 mm long). Shoot-producing cultures were obtained from 4, 32 and 70% of meristem lips, shoot tips and nodal segments within 12, 6 and 4 weeks, respectively. The genotype of cultures had a greater influence on development of shoot-producing cultures than medium composition. Cultivars Max/Ber and Oxford had the highest rates of establishment in culture and subsequent shoot proliferation, while P. tacamahaca, P. trichocarpa and cv. Robusta exhibited very low rates of establishment and low vigor in vitro. Shoot tip development was best on agar-solidified medium whereas liquid medium resulted in vitrification. Higher rates of axillary shoot production from established cultures were obtained with benzyladeninc or zeutin than with 2-isopen-tenyladenine. deducting the benzyladenine concentration from 4,4 to 1.1 μ M , increased the production of elongated shoots suitable for rooting.     

A method for long-term micropropagation of Phaseolus coccineus L.

A method for long-term plant regeneration of Phaseolus coccineus L, is described. Shoot-tips and cotyledonary nodes cultured on a Murashige and Skoog medium supplemented with N⁶-benzylaminopurine, 10 M, and -naphthaleneacetic acid, 1M, formed multiple bud-shoots. These shoots were transferred to medium containing BAP 1 M, NAA 0.1 M, and gibberellic acid 3 M to promote shoot growth and further shoot multiplication. Rooting was achieved in medium with 11 M indole-3-acetic acid. Rooted plants grew to maturity and were fertile. Cultures have maintained their ability to regenerate plants for more than two years. A sample of 30 regenerated plants (R₀) was tested for chromosome number, all of them being diploid; seven isozymatic systems were electrophpretically analyzed in 82 R₀ regenerated plants. No differences were observed in their electrophoretic patterns in comparison with those shown by seedlings. Histological studies revealed the origin of buds from calluses via organogenesis.Abbreviations BAP N⁶-benzylaminopurine - 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid - MS Murashige and Skoog (1962) medium - NAA -naphthaleneacetic acid - ADH alcohol dehydrogenase - GOT glutamic-oxaloacetic transaminase - MDH malate dehydrogenase - 6PGD 6-phosphogluconate dehydrogenase - PGI Phosphoglucose isomerase - PGM phosphoglucose mutase - SK shikimate dehydrogenase     

A scanning electron microscope study of normal and vitrified leaves from Datura insignis plantlets cultured in vitro

The surface anatomy of normal and vitreous leaves of plantlets obtained from Datura insignis Barb Rodr nodal segment cultures was compared using scanning electron microscopy. Normal and vitrified leaves are similar in several ways. They are both amphistomatic, and have similar distributions of glandular and non-glandular trichomes. Stomata have similar length, diameter and distribution in normal and vitreous plants. Immature stomata, which have closed pores, and plugged stomata, which contain an amorphous material between their guard cells, occur in both normal and vitrified leaves. Normal and vitreous leaves differ in the frequency of normal and abnormal stomata. Normal stomata have kidney-shaped guard cells and resemble closely those found in field-grown plants, whereas abnormal stomata have deformed guard cells. Normal stomata represent approximately 80% of the total number of stomata in normal leaves, but only 7% of the total number of stomata in vitreous leaves. Abnormal stomata represent 90% of the total number in vitreous leaves. The deformation of guard cells could possibly be a mechanical impediment to stomatal function.     

Development of a new culture medium and efficient protocol for in vitro micropropagation of Ceratonia siliqua L.

A new basal culture medium was developed and tested using a rapid and efficient protocol of in vitro axillary shoot bud proliferation of Ceratonia siliqua L., an important Mediterranean Fabaceae plant species. In a first experiment, the new formulated ‘LA’ mineral composition significantly improved shoot growth and proliferation as compared with Murashige and Skoog medium (MS, 1962) in both solid and liquid culture media. However, the liquid culture system proved to be the most suitable for shoot induction, shoot length (about fourfold higher), and multiplication rate (about two-fold higher), the difference being significant. The measured growth and proliferation parameters were further improved when LA mineral composition was optimized, in a second experiment. The highest multiplication rate (6.3) was achieved during the second subculture using the optimized ‘LAC’ medium. Noticeably, hyperhydricity and shoot-tip necrosis symptoms were absent in both formulated LA and LAC compositions when using the liquid culture system. In vitro rooting in solid medium showed 41.7 to 46.3% response on a solid medium which was more suitable than the liquid culture system, the difference being significant. In contrast, pretreated microcuttings with 3 μM IBA (indole-3-butyric acid) were successfully rooted ex vitro, showing significantly higher response (91.7%), average root number (8.3), and root length (31.5 mm). The plantlets were successfully acclimatized showing more than 90% survivability and normal morphology. The present study is a first cost-effective protocol for carob micropropagation combining the use of the newly formulated LAC basal medium, a liquid culture system, and ex vitro rooting.

     

A role of microtubules in the polarity of statocytes from roots of Lepidium sativum L.

When roots of Lepidium sativum L. are immersed in a colchicine solution (10^-4 mol l^-1), the cortical microtubules of statocytes are affected such that the dense network ofmicrotubules at the distal cell edges, between the endoplasmic reticulum and the plasma membrane, disappears almost completely, whereas the microtubules, lining the anticlinal cell walls are reduced only to a limited extent. Upon inversion of colchicine-pretreated roots, the distal complex of endoplasmic reticulum sinks into the interior of the statocyte. Germination of seeds in the cold (3–4°C) leads to a retardation of statocyte development; the elaborated system of endoplasmic reticulum is lacking, and only a few microtubules are observable, lining the plasma membrane along the anticlinal cell walls. During an additional 4 h at 24°C, groups of microtubules develop near the plasma membrane in the distal one-third of the statocytes, coaligning with newly synthesized cisternae of the endoplasmic reticulum. It is proposed that, particularly at the distal statocyte pole, microtubules in coordination with cross-bridging structures, act in stabilizing the polar arrangement of the distal endoplasmic reticulum and, in turn, facilitate an integrated function of amyloplasts, endoplasmic reticulum and plasma membrane in graviperception.Abbreviations ER endoplasmic reticulum - MT microtubule     

Large-scale production of Anogeissus pendula and A. Latifolia by micropropagation

Summary Success has been achieved in developing a complete protocol for mass propagation of Anogeissus pendula and A. latifolia, two important forest species found in India. Seeds cultured on plant growth regulator-free, semisolid Murashige and Skoog (MS) medium germinated within 5–6 wk and formed 4–6-cm long shoots. The shoots multiplied on MS+4.4 μM benzyladenine (BA)+5.7 μM indoleacetic acid (IAA) + casein hydrolysate (100 mgl⁻¹) + ascorbic acid (50 mgl⁻¹) + sucrose (3%) + agar (0.8%). A majority of the genotypes rooted with more than 90% efficiency when 5–6 cm individual shoots were cultured on 1/2MS (only major salts reduced to half strength)+2.3 μM IAA+2.5 μM indolebutyric acid (IBA) + sucrose (3%)+agar (0.8%) for 15 d. Those 10% (approx.) genotypes that did not root well on the above medium could be rooted with ease by increasing the concentration of IAA in the rooting media from 2.3 to 5.7 μM. The in vitro-raised plants were successfully transferred to the soil with a success rate of over 85%. Using this protocol, over 560 000 tissue-cultured plants of these two species have been produced and dispatched to various state forest departments for field trials and routine plantations.     

Propagated fluctuations of the electric potential in the apoplasm of Lepidium sativum L. roots

The electric potential on the surface of the Lepidium sativum L. root apex was recorded by means of six non-polarizable electrodes. Nonevoked fluctuations of the potential with amplitudes below 0.1 mV were observed. The fluctuations could be reversibly inhibited either by ether vapor or by anoxia caused by N₂. They did not occur in killed roots. Cross-correlation analysis of the fluctuations from six electrodes located one above another along the 3-mm apical region showed a pattern of time delay which indicates that the fluctuations may be the consequence of signals propagated in the root with a velocity of 3–9 mm · s^–1 in a basipetal direction from the root cap. We hypothesize that the fluctuations are due to signals of an unknown nature propagated along an intrasymplasmic continuous system, the symreticulum, composed of the cortical ER of individual cells and desmotubules passing through the plasmodesmata.Abbreviations AC alternating current - AP action potential - ACF autocorrelation function - CCF cross-correlation function - DC direct current - EEP extracellular electric potential This research was supported by Bundesminister für Forschung und Technologie, Bonn, and Ministerium für Wissenschaft und Forschung, Düsseldorf, (AGRAVIS). We are grateful to Mr. Dipl.-Ing. P. Blasczyk for constructing the amplifiers and for advice in instrumentation, and to Mr. H. Laubach for constructing the mechanical assembly.     

大白菜原生质体培养再生体系的优化

以栽培大白菜亲本材料河头早A、河头早B、青麻叶C、青麻叶D、石特一号9-3、石特一号11-4、玉青、运农一号等8个品种的无菌苗下胚轴为外植体游离获得原生质体,用液体浅层法将原生质体培养在含1．5mg,NAA、0．5mg／L TDZ、0．6mg／LBA、0．5mg／L 2,4-D的DPD培养基中。培养约48h,细胞发生第一次分裂。培养到第6天发生第二次分裂。约4周,形成大量细胞团和肉眼可见的小愈伤组织。当愈伤组织长到直径4mm左右时,转入分化培养基中诱导分化,在含0．05mg／L IBA的培养基中诱导生根。对影响原生质体培养与分化的多种因素进行了较为系统的研究。     

Invasion of Lepidium draba (Brassicaceae) in the western United States: distributions and origins of chloroplast DNA haplotypes

Advances in phylogeography are of great value for understanding the population structure and origins of invasive genotypes. Such insights provide constructive information for current or future biological control research efforts. In this study, we investigated a highly variable chloroplast DNA (cpDNA) marker for populations of the weed Lepidium draba (Brassicaceae) in its native Eurasian and invasive US ranges. We sequenced DNA from 684 individuals from Eurasia and the US and found 41 different haplotypes. Our comparative study between the native and invasive ranges showed a 33% reduction in allelic richness (A) and a 7% reduction in haplotype diversity (h) since introduction into the US. Most genetic variation in the native range was observed within geographical regions and populations, not between regions, and this result was similar for the invasive range. Assignment tests indicated the most likely origins of many invasive haplotypes. Some of these occurred in western Europe, supporting an expanded native range that had been proposed for the species. Exact locations were identified for a diverse set of invasive haplotypes which can be used in ongoing host-specificity tests of potential biological control agents.     

Bud regeneration from inflorescence explants for rapid propagation of geophytes in vitro

Bulbs, corms and other subterranean storage organs are commonly used as explant source material for the establishment of geophytes in vitro. The inflorescence stalk was found to be a good alternative source of explants to overcome explant contamination originating from underground storage organs. Inflorescence explants of Allium, Dichelostemma, Eucrosia, Gladiolus, Haemanthus, Hyacinthus, Narcissus, Nerine and Ornithogalum were used to establish cultures in vitro. The regeneration potential of the inflorescence was compared with regeneration from bulb twin scales or from apical buds isolated from corms. Gladiolus (Iridaceae) explants isolated from the floral stem just below the expanding florets, still enclosed in the bracts, were highly regenerative in the presence of naphthalene acetic acid (NAA) and kinetin. In the presence of 2,4-dichlorophenoxyacetic acid and benzyl aminopurine (BA) in the medium, explants isolated from the tissue at the junction between the peduncle and the pedicels of a young Nerine (Amaryllidaceae) inflorescence regenerated several buds. The scapes of young unemerged inflorescences taken from sprouting bulbs of Narcissus (Amaryllidaceae), following a 15 °C storage treatment, regenerated buds in the presence of NAA, BA, elevated phosphate and adenine sulfate in the medium. The number of buds regenerated depended on the location on the scape from which the explant was isolated, and on the duration of the 15°C treatment. In Allium (Alliaceae), capitulum tissue between the flower pedicels regenerated buds. Explants excised from the peduncle, as well as the pedicel-peduncle junction of Dichelostemma (Alliaceae), Ornithogalum, Hyacinthus (Hyacinthaceae) and Eucrosia (Amaryllidaceae) regenerated several buds in each type of explant. In the case of Haemanthus (Amaryllidaceae), pedicel-peduncle junction explants regenerated buds only when excised from inner whorl florets. Propagation protocols and the potential use of expediently isolated inflorescence explants for efficient micropropagation of geophytes are discussed. Received: 1 September 1999 / Revision received: 13 December 1999 / Accepted: 13 December 1999     

Relationship between Linear Growth and the Distance of Initiation of Lignification behind the Root-tip in Lepidium sativum L.

The relationship between the rate of linear growth and distanceof initiation of the zone of lignification behind the root-tipin young roots of Lepidium sativum L. was investigated underdifferent cultural conditions. The distance varied between 1.0 mm and 3.4 mm. All the factorswhich retard linear growth reduce the distance of the zone oflignification behind the root-tip, and an acceleration in lineargrowth increases the distance correspondingly. The reductionor extension of this distance depends upon the intensity oflinear growth, and is independent of the nature of the inhibitingagent employed. The factors and substances used were temperature,mannitol, respiratory inhibitors like KCN, 2-4 DNP, sodium fluorideand p-nitrophenol IAA, coumarin, and nitrate and phosphate.     

A rapid protocol for the authentication of isolated differential display RT-PCR CDNAs.

The most time-consuming and problematic step in the overall DDRT-PCR technique is the confirmation that the isolated cDNA clone represents a differentially expressed gene. We have previously suggested that the majority of apparent false positives generated by DDRT-PCR do in fact result from the PCR reamplification of cDNA species which co-migrate with the cDNA of interest, and we have outlined a procedure to effectively eliminate these from further study. However, in situations where RNA is limiting, it is still desirable to confirm that a purified cDNA amplicon does, in fact, represent the originally observed differentially expressed gene prior to embarking on expression studies.     

An efficient protocol for micropropagation of old cypress of Abarkuh (Cupressus sempervirens var. horizontalis [Mill.]) under in vitro condition

Plant Cell, Tissue and Organ Culture (PCTOC) - This study was carried out to investigate the DNA markers between diploid and colchicine-induced autotetraploid Platycodon grandiflorum A. De Candolle...     

Optimisation of a rapid and efficient transformation protocol for fungal blast-susceptible indica rice cultivars HR-12 and CO-39

Rice is an important staple crop and fungal blast disease destroys about 10–30% of its global produce, annually. Although genetic manipulation has largely been employed in crop-improvement programmes and agricultural biotechnology, the ease of transformation of several recalcitrant indica cultivars continues to be a challenge. HR-12 and CO-39 are two indica cultivars that are commonly used in breeding programmes, but are susceptible to biotic threats like fungal blast and sheath blight disease. Here in this study, we have optimised a rapid and reproducible transformation protocol for the said cultivars, having compared both the tissue-culture and in-planta methods of transformation. Murashige & Skoog basal media supplemented with maltose and 2.5 mg l⁻¹ 2,4-D induced efficient callogenesis in HR-12, while maltose with 3 mg l⁻¹ 2,4-D gave optimum results in case of CO-39. The media containing 0.5 mg l⁻¹ NAA, 3 mg l⁻¹ BAP, and 1 mg l⁻¹ kinetin yielded a maximum regeneration efficiency of 62% and 65% in HR-12 and CO-39, respectively. The studies with Agrobacterium tumefaciens, LBA4404 strain harbouring pCAMBIA1303 suggested that although these cultivars demonstrated successful gene-transfer, they failed to regenerate efficiently, post-transformation. Alternatively, our modified in-planta piercing and vacuum infiltration-based protocol resulted in 33–35% transformation efficiency in less than half the time required for tissue-culture based transformation method. As per our knowledge, it is among the highest obtained from existing piercing-based direct transformation protocols in rice, and can also be implemented in genetically manipulating other recalcitrant varieties of rice.     

Inheritance of seed yield and quality traits in peas (Pisum sativum L.)

Summary A half diallel analysis involving nine cultivars showed that additive as well as non-additive gene effects were important for the inheritance of seed yield per plant, 100-seed weights, protein content and potassium per cent. For remaining traits non-additive genetic components were important. Overdominance was observed for all traits except for 100-seed weight, which expressed partial dominance. Parents PMR-T10, EC21857, EC109182, T163 and EC109189 were good general combiners for seed yield, seed weight and quality traits. In general there was a good relationship between per se performance and the gca effects of the parents for all traits. Cross combinations such as LMR8 x EC109182,LMR8 x PMR-T10,LMR8 x EC21857,PMRT10 x EC21857 and P23 x EC21857 were found promising. The seed yield was positively correlated with other quality traits. Protein had a positive correlation with methionine and phosphorus. All the values of correlation co-efficients were non-significant except for yield with potassium, 100-seed weight and protein with methionine, indicating that yield and quality attributes can be improved simultaneously by simple selection procedures.