Ethanol synthesis, nitrogen, carbohydrates, and growth in tissues from nitrogen fertilized Pseudotsuga menziesii (Mirb.) Franco and Pinus ponderosa Dougl. ex Laws. seedlings

 Seedlings of Douglas-fir, Pseudotsuga menziesii (Mirb.) Franco, and ponderosa pine, Pinus ponderosa Dougl. ex Laws., were grown in a controlled environment and fertilized with nutrient solutions containing 150 ppm (+N), or 0 ppm nitrogen (−N). These treatments greatly altered seedling growth, and the concentrations of N and carbohydrates in their tissues. Metabolically active tissues, such as roots, incubated with a limited supply of O₂ became hypoxic faster and synthesized more ethanol than less active tissues, such as needles. All tissues that were incubated for 4 h in N₂ synthesized ethanol. Needles incubated in N₂ and light had much lower quantities of ethanol than needles in N₂ and dark, suggesting that O₂ from photosynthetsis limited internal anoxia. Most tissues from +N seedlings synthesized greater quantities of ethanol in N₂ anoxia than tissues from −N seedlings, probably because they were able to produce more enzymes with a greater availability of N. However, this increase in ethanol synthesis between N treatments was most pronounced in the phloem. Ethanol and soluble sugar concentrations were negatively related in needles and positively related in roots of N+ seedlings, but not −N seedlings. Starch concentrations had no effect on the amount of ethanol produced by any tissue. Regardless of N treatments, all tissues from ponderosa pine produced more N₂-induced ethanol than Douglas-fir, in part because its tissues contained different concentrations of soluble sugars and N as a consequence of phenological differences between the species. However, ponderosa pine tissues may also maintain greater quantities of anaerobic enzymes, or their isozymes than Douglas-fir. Received: 22 February 1998 / Accepted 23 June 1998     

In vitro differentiation of plantlets from embryonic explants of lodgepole pine (Pinus contorta Dougl. ex Loud.)

A protocol is described for plantlet formation in juvenile tissues of Pinus contorta. Shoots were induced on embryonic, cotyledonary and hypocotyl explants cultured on a defined medium supplemented with cytokinin. The concentration of salts, vitamins and cytokinin (benzylamino purine) in the medium, as well as different temperature regimes, strongly influenced the frequency of bud formation. Differentiation of shoot primordia and their subsequent development was also markedly affected by cytokinin exposure times. Bud development and elongation were enhanced by elimination of the phytohormone, reducing the strength of mineral salts, vitamins and sucrose in the medium, as well as by the inclusion of charcoal. Rooting was induced by treating the shoots with a sterilized rooting powder containing indole-butyric acid and culturing them in agar-solidified medium containing reduced mineral salts, vitamins, sucrose and charcoal. The number of chromosomes and their structure were found to be normal in the regenerated plantlets.     

Effect of moisture stress on the growth ofPinus ponderosa Dougl. ex. Laws seedlings in relation to their field performance

Forty wind-pollinated families from eight provenances, four from xeric and four from mesic site types in the Sierra Nevada of California, were examined in the greenhouse for differential response to soil moisture stress. Analysis was made of shoot-root allometric growth coefficients, absolute and relative growth rates and the relationship among these response measures and growth in three field plantations. The results showed significant differences among families for allometric growth (k) coefficients under moderate stress, but no differences among provenances or between site types. Absolute and relative growth rate differences between site types were not significant under any treatment; however provenance differences were observed. Correlation coefficients between plantation performance and greenhouse growth estimates were sometimes significant but significance was plantation and treatment specific. Research performed while author was on sabbatical leave at the Institute of Forest Genetics, USDA-USFS, Placerville, California.     

Influence of ovule perforation,plant growth regulators,andl-glutamine on in vitro growth of immature peach embryos

Summary Ovule perforation technique and media components (plant growth regulators andl-glutamine) were tested on in vitro growth of immature (<3 mm) embryos of “Springcrest” and “Earligrande” peaches. Ovule perforation was 2 to 4 times more effective in promoting embryo growth than leaving ovules intact.l-Glutamine (400 mg·liter⁻¹) promoted an increase in growth but could not be used with indole-acetic acid plus kinetin because an antagonistic effect on embryo growth occurred. The use of these exogenous plant growth regulators did not increase embryo growth over in vivo growth.     

Effect of plant growth regulators on in vitro fiber development from unfertilized and fertilized Egyptian cotton ovules

Unfertilized and fertilized ovules of Gossypium barbadense Giza 45 (extra long staple variety) were used to study the effect of plant growth substances (auxins, gibberellins and cytokinins) on in vitro fiber initiation and development. Kinetin, alone did not increase total fiber unit (TFU) of unfertilized ovules, while an increase in TFU value occurred when a constant level of IAA and GA₃ were used either separately or in combination in the liquid medium. GA₃ used alone, produced a higher TFU value than that produced by IAA, whilst, IAA with a constant level of GA₃ (5 M) produced the highest value of TFU. GA₃ with a constant level of IAA (5 M) produced a lower TFU value. Kinetin reduced the stimulatory effect of IAA and GA₃ on TFU value when used in combination with either substance. In fertilized ovules, the highest level of TFU was reached when IAA, with a constant level of GA₃, was added to the medium, whilst its lowest level was obtained when IAA was used alone. Estimation of in vitro fiber production, as well as the effect of growth substances used in different concentrations on in vitro fiber initiation and development from unfertilized and fertilized ovules of Egyptian cotton varieties Gossypium barbadense Giza 45 are discussed.     

Influence of growth regulators on plant regeneration from epicotyl and hypocotyl cultures of two groundnut (Arachis hypogaea L.) cultivars

This study was designed to evaluate the effect of phytohormones on plant regeneration from epicotyl and hypocotyl explants of two groundnut (Arachis hypogaea) cultivars. Explants cultured on media with auxins and in combination with cytokinin produced high frequency of callus. After four weeks, callus from these cultures was transferred to medium with cytokinin and reduced auxin, shoot buds regenerated from the cultures. A high rate of shoot bud regeneration was observed on medium supplemented with 2.0 mg/L BAP and 0.5 mg/L NAA. Among the different auxins tested, NAA was found to be most effective, producing the highest frequency of shoot buds per responding cultures. Of the two explants tested, epicotyl was found to be best for high frequency shoot bud regeneration. Multiple shoots arose on MS medium supplemented with BAP or kinetin (1.0–5.0 mg/L) plus IBA (1.0 mg/L), with maximum production occurring at 5.0 mg/L. The elongated shoots developed rootsin vitro upon transfer to MS medium supplemented with NAA or IBA (0.5–2.0 mg/L) and kinetin (0.5 mg/L) for 15 days.In vitro produced plantlets, were transferred to soil and placed in a glasshouse developed successfully, matured, and set seeds.     

Effect of plant growth regulators on the cellular growth and levels of intracellular protein,starch and polyamines in embryogenic suspension cultures of Pinus taeda

Somatic embryogenesis is the most important in vitro culture system for conifer propagation. However, Pinus taeda has been considered recalcitrant to somatic embryogenesis in commercial scale-up. The study of biochemical and physiological aspects of cell growth could lead to a better understanding of somatic embryogenesis in this species. In the present work, we investigated the cell growth dynamics, intracellular levels of proteins, starch and polyamines in suspension cultures of Pinus taeda established in plant growth regulator-free medium (BM0) and in medium supplemented with 2 M 2,4-dichlorophenoxyacetic acid, 0.5 M 6-benzylaminopurine and 0.5 M Kinetin (BM2). Cell cultures growing in BM0 medium showed an increase in the sedimented cell volume from 3.77 to 17.73 ml after 24 days of culture. Those cultured in BM2 medium showed an increase in the sedimented cell volume from 4.23 to 25.17 ml after 20 days of culture. Intracellular proteins levels increased during the exponential growth phase and starch levels decreased until the exponential phase, followed by a synthesis up to the stationary phase, in both BM0 and BM2 media. Highest putrescine levels occurred in cultures growing in BM0 medium and this was associated with the low cellular growth.     

Effect of exogenous plant growth regulators on in vitro seed growth,embryo development,and haploid production in a cross between H. vulgare (L.) and H. bulbosum (L.)

Exogenous plant growth regulators are known to increase the efficiency of interspecific and intergeneric crosses. In vitro floret culture provides a defined system for assessing the importance of various plant growth regulators on the determinants of haploid production efficiency (seed set, embryos per seeds, and plants per embryos) in Hordeum vulgare × Hordeum bulbosum crosses. The individual and combined effects of three plant growth regulators (2,4-D, GA₃ and kinetin) on in vitro seed growth, embryo development and haploid production efficiency were tested in floret culture of the cross H. vulgare, cultivar Klages × H. bulbosum. All treatments, except kinetin alone, produced larger seeds and more embryos/100 seeds than the control (no plant growth regulator). 2,4-D alone was superior to GA₃ alone in haploid production efficiency (70.6 vs. 51.5) as measured by the number of plants regenerated/100 florets pollinated. Although kinetin +2,4-D+GA₃ produced the largest seeds and embryos, no advantage over 2,4-D alone was observed in haploid production efficiency. 2,4-D alone or kinetin +2,4-D are recommended for the purpose of barley haploid production in floret culture using the bulbosum method.Abbreviations 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid     

Influence of some plant growth regulators on the growth and organogenesis ofCymbidium aloifolium (L.) Sw. seed-derived rhizomesin vitro

Summary An efficient procedure is outlined forin vitro regeneration of an epiphytic orchid,Cymbidium aloifolium (L.) Sw. using rhizomes developed from seeds. Murashige and Skoog's (1962) medium (MS) containing indole-3-acetic acid (IAA), indole-3-butyric acid (IBA), or 1-naphthaleneacetic acid (NAA) stimulated growth and proliferation of rhizomes with NAA being most effective at 5.0 mg.l⁻¹ (27.0 μM). Shoot bud differentiation was induced in the apical portions of the rhizomes on MS medium containing kinetin (Kn) or N⁶-benzyladenine (BA). The highest frequency of shoot regeneration (91.5%) and the maximum number of shoot buds formed (3.5 shoots/rhizome) were recorded with BA at 1.0 mg.l⁻¹ (4.4 μM). NAA (0.1 mg.l⁻¹, 0.54 μM), whenever added to the medium in conjunction with BA (1.0 mg.l⁻¹, 4.4 μM), slightly enhanced the frequency of shoot bud regeneration (92.6%) and the number of shoot buds formed (5.2 shoots/rhizome). Moreover, an NAA-BA combination induced rooting in regenerated shoots thereby producing complete plantlets in one step. Shoots developed on cytokinin-supplemented medium were rooted on MS containing NAA at 1.0 mg.l⁻¹ (5.4 μM). Regenerated plantlets were acclimated and eventually established in a garden.     

Distinguishing the effects of light and temperature variations on the growth, development, multiplication potential and ex vitro survival rates of in vitro cassava

Dissemination of cassava tissue culture plantlets is difficult in the arid tropics due to low eux vitro survival rates. Increased in vitro light intensity has been reported to induce high ex vitro survival rates. The results from earlier experiments suggested that it would be worthwhile to analyse the separate effects of in vitro light and of temperature on the in vitro growth pattern as well as differentiate its relation to ex vitro survival. Accordingly, analysis of a range of in vitro light intensities from 0 to 369 μmol^?1 m^?2 photosynthetic photon flux density (PPFD) was conducted both with and without fans to control the heat. Temperature proved stable at low PPFD levels but increased above 204 μmol s^?1 m^?2 when no fans were used. Increased PPFD levels induced larger fresh and dry masses as well as stem thickness. PPFD levels affected the developmental index (senescent leaf numbers) in vitro when it rose above 204 μmol s^?1 m^?2 PPFD. Raised temperature ranges increased the multiplication index (node numbers) in vitro and ex vitro. It increased root number and leaf development (lobe anatomy). As in vitro temperatures of up to 40°C improve multiplication rates and PPFD levels above 101 μmol s^?1 m^?2 were detrimental for ex vitro survival (as low as 60%), it is suggested that simpler and less costly laboratories with low light levels and a wide range of temperature tolerance could be successfully established in the tropics for in vitro cleaning and rapidly multiplying crops like cassava.     

Effect of basal medium, growth regulators and Phytagel concentration on initiation of embryogenic cultures from immature zygotic embryos of loblolly pine (Pinus taeda L.)

Low initiation frequency is one of the main barriers in applying somatic embryogenesis to the clonal production of Pinus species. Factors affecting initiation, including basal medium, plant growth regulators, and Phytagel concentration, have been investigated in loblolly pine (Pinus taeda L.). BM₁ basal medium proved superior to DCR₁ and LP (LP basal salts plus BM₁ organic nutrients). No extrusion from megagametophytes was exhibited on LP medium. The combination of 3 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D) and 0.5 mg/l 6-benzylaminopurine (BA) resulted in a higher extrusion frequency than that of 11 mg/l 2,4-D, 4.5 mg/l BA and 4.3 mg/l kinetin. Phytagel at 1 g/l resulted in the highest explant browning, but the lowest extrusion frequency, while 4 g/l Phytagel induced some dry embryogenic extrusions. Phytagel at 2 g/l was regarded as the best level for initiation of embryogenic cultures. Received: 23 December 1996 / Revision received: 22 July 1997 / Accepted: 2 September 1997     

不同生长调节剂对狗肝菜愈伤组织诱导和离体快繁的影响

目的 :研究不同生长调节剂对狗肝菜愈伤组织诱导和离体快繁的影响。方法 :狗肝菜不同外植体在附加不同生长调节剂的培养基上诱导愈伤组织 ,比较愈伤组织的诱导率 ;用 3因子 5水平的正交实验 ,比较不同生长调节剂对丛生芽诱导的影响 ;在附加不同生长调节剂的培养基上比较芽增殖倍数 ;附加不同浓度NAA的培养基上比较生根效果。结果 :愈伤组织诱导率相对以叶片最高 ,茎段次之 ,最后为叶柄 ;愈伤组织诱导的最佳培养基为MS 6-BA0 .5 NAA1 .5 ;不同激素对茎段芽诱导的影响次序为 6-BA>KT >NAA ,芽诱导的最佳培养基为MS 6-BA2mg/L KT1mg/L NAA0 .5mg/L ;芽继代增殖的最佳激素组合是MS 6-BA2mg/L NAA2mg/L ,增殖倍数达 3.0 0 ,影响芽继代增殖的因素次序为 6-BA >NAANAA0 .5mg/L的生根效果较好。结论 :附加一定的生长调节剂能提高狗肝菜愈伤组织的诱导率和离体快繁的效率。     

Development of the interferon system. I. In chicken cells development in ovo continues on time in vitro

Summary When confluent monolayers of cells derived from chicken embryos of different gestational age were cultured for several days without a medium change, a condition termed in vitro aging, the cells' developed an increased capacity to express the interferon (IFN) system. The capacity to both produce IFN and to respond to its antiviral action were enhanced up to 1000- and 100-fold, respectively. Remarkably, the programmed development of the IFN system in these cells seemed to continue virtually uninterrupted after monodispersion of the cells and seeding at high cell density. Cells prepared from young embryos required more time to develop the IFN system than cells from older embryos with the yield of IFN, and sensitivity to its action, related directly to the total in ovo and in vitro age of the cells in culture. For example, essentially the same yields of IFN were obtained from cell cultures made from 5-d-old embryos “aged” for 10 d in vitro, as were obtained from 10-d-old embryos whose cells were aged in vitro for 5 d. In contrast, inducibility of 2′–5′ oligoadenylate synthetase by IFN and the induction of heat shock genes by elevated temperature are not enhanced with in vitro aging. The programmed development of the IFN system that starts in ovo seems to continue on schedule in vitro, making the development of the IFN system in chick embryo cells appear as a time-dependent process. This study was supported by the grant RO1 AI18381 from the national Institute of Allergy and Infectious Diseases, Bethesda, MD, and benefited from services of the Cell Culture Facility of the Biotechnology Center at The Univeristy of Connecticut.     

Influence of plant growth regulators, polyamines and glycerol interaction on growth and morphogenesis of carposporelings of Grateloupia cultured in vitro

The influence of the plant growth regulators 2,4-D, GA₃, BA and kinetin, and the polyamines putrescine, spermidine and spermine were tested on axenic in vitro cultures of carposporelings of Grateloupia doryphora. The auxin 2,4-D (10^-3 M) and the polyamine spermine (10^-6 M and 10^-3 M) induced a callus (disorganised cell mass that arose from the organised tissue of the carposporeling, as demonstrated by microscopic monitoring of the tissue). Putrescine and spermidine (10^-3 M) transformed the carposporelings into cell masses that produced shoots. BA (10^-3 M) and kinetin (10^-6 M and 10^-3 M) were inhibitory. In 10^-1 M glycerol-containing culture medium, which is known to induce the formation of morphogenic cell masses, the addition of GA₃ M) resulted in the inhibition of the morphogenesis (i.e. shoot emission) in the cell mass. The kinetin at 10^-6 M inhibited morphogenesis, whilst at 10^-3 M inhibited even the formation of the cell masses. The combination of glycerol (10^-1 M) and the auxin 2,4-D (10^-6 and 10^-3 M) or the polyamines putrescine, spermidine and spermine (10^-6 and 10^-3 M) resulted in a bigger size of the cell masses that led to a higher amount of shoots per cell mass than in glycerol alone. This revised version was published online in June 2006 with corrections to the Cover Date.     

Influence of plant growth regulators on micropropagation and in vitro flowering of Trichodesma indicum (Linn) R. Br

An efficient protocol for micropropagation and in vitro flowering of Trichodesma indicum (Linn) R. Br. was developed using shoot tip explants. The physiological role of cytokinin and its combination with auxins on micropropagation and in vitro flowering was investigated. The highest number of shoots (9.94 ± 0.10) and the maximum average shoot length (5.56 ± 0.35 cm) were recorded on Murashige and Skoog (MS) medium supplemented with benzylaminopurine (BAP) (4.44 μM) and naphthaleneacetic acid (NAA) (2.69 μM). The effect of sucrose concentration on in vitro floral development was studied in plantlets cultured on MS medium supplemented with gibberellic acid (GA₃) and BAP. The highest percentage of flowering (93.2%) was obtained on MS medium supplemented with GA₃ (1.44 μM), BAP (1.33 μM) and sucrose (30 g l^?1). Root formation from the adventitious shoots was easily achieved on MS medium containing indole-3-butyric acid (IBA) (2.46 μM). The regenerated plantlets showed 86% survival rate and were phenotypically normal. The described method can be successfully employed for large-scale multiplication and in vitro flowering of T. indicum.     

Influence of growth regulators on the development,quality, and physiological state of in vitro-propagated Lamprocapnos spectabilis (L.) Fukuhara

In Vitro Cellular & Developmental Biology - Plant - There is little information on the in vitro tissue culture systems in Lamprocapnos spectabilis (bleeding heart). The aim of this study was to...     

Components of the gaseous environment and their effects on plant growth and development in vitro

This article has first been published in: P. J. Lumsden, J. R. Nicholas and W. J. Davies (eds.), Physiology, Growth and Development of Plants in Culture, 165–190, 1994.     

Influence of carbon source and concentration on the in vitro development of olive zygotic embryos and explants raised from them

The influence of sucrose or mannitol on in vitro zygotic embryo germination, seedling development and explant propagation of olive tree (Olea europaea L.) was compared. Embryos germinated without sucrose in the medium but for adequate development of the seedlings to yield viable plants, a carbohydrate supply was necessary; both sucrose and mannitol were equally suitable for this purpose. However, when explants obtained from in vitro germinated embryos were cultured with mannitol or sucrose, then the polyalcohol promoted significantly more growth than sucrose by increasing shoot length, pairs of leaves formed, and breaking apical dominance. This improved the in vitro culture of olive plant material, thus allowing new olive clonal lines to be obtained in shorter times. This will assist in future breeding experiments with the species.     

Light spectral quality effects on the growth of potato (Solanum tuberosum L.) nodal cuttings in vitro

Summary The effects of light spectral quality on the growth of in vitro nodal cuttings of potato (Solanum tuberosum L.) cultivars Norland, Superior, Kennebec, and Denali were examined. The different light spectra were provided by Vita-Lite fluorescent (VF) (a white light control), blue fluorescent (BF), red fluorescent (RF), low-pressure sodium (LPS), and a combination of low-pressure sodium plus cool-white fluorescent lamps (LPS/CWF). For all cultivars, stem lengths after 4 wk were longest under LPS, followed by RF, LPS/CWF, VF, and BF (in descending order). Microscopic studies revealed that cells were shortest when cultured in BF or VF environments, and were longest in RF or LPS lamp environments. The highest number of axillary branches occurred on plantlets grown with LPS or LPS/CWF, whereas the lowest number occurred with BF. No leaf or stem edema (callus or gall-like growths) occurred with LPS or LPS/CWF lighting, and no edema occurred on cv. Norland plantlets, regardless of lighting. Results suggest that shoot morphologic development of in vitro grown potato plants can be controlled by controlling irradiant spectral quality.