Effect of plant growth regulators on ethylene production, 1-aminocyclopropane-1-carboxylic acid oxidase activity, and initiation of inflorescence development of pineapple

With the development of pineapple [Ananas comosus (L.) Merr.] as a fresh fruit crop, it became common to force inflorescence development with ethephon [(2-chloroethyl)phosphonic acid] or ethylene throughout the year. Environmental induction (EI) of inflorescence development disrupts scheduling of fruit harvest and may cause significant losses if small plants are induced, resulting in fruits that are too small to be marketable. Our objective was to identify plant growth regulators (PGRs) that could inhibit EI. Because circumstantial evidence indicates that EI occurs in response to naturally produced ethylene or changes in plant sensitivity to it, most work was done with PGRs that inhibit ethylene biosynthesis or block ethylene action. The synthetic auxin 2-(3-chlorophenoxy)propionic acid (CPA) was included because in one study it reduced the percentage of EI. GA₃, aminooxyacetic acid (AOA), aminoethoxyvinylglycine (AVG), daminozide [butanedioic acid mono-(2,2-dimethylhydrazide)], and silver thiosulfate (STS) had no effect on EL CPA, paclobutrazol [(2RS,3RS)-1-(4-chlorophenyl)methyl-4,4-dimethyl-2(1h-1,2,4-triazol-1-yl)penten-3-ol], and uniconazole [(E)-(p-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-penten-3-ol] delayed or inhibited EI of pot-grown pineapple plants. Uniconazole and paclobutrazol inhibited growth and ethylene production by leaf basal-white tissue, and either or both effects could account for the inhibition of EI. Production of 1-aminocyclopropane-1-carboxylic acid (ACC) was unaffected by these compounds, but the activity of ACC oxidase, which converts ACC to ethylene, was inhibited and probably accounts for the reduced ethylene production by leaf basal-white tissue. CPA stimulated ethylene production by stem apical tissue approximately fourfold relative to the control. ACC oxidase activity and the malonyl-ACC (MACC) content in stem apical tissue were also greater than in the control, indicating that CPA greatly stimulated the production of ACC and its sequestration into MACC. The mechanism by which CPA delayed or inhibited EI is not known. CPA, paclobutrazol, and uniconazole appear to have some potential for inhibiting EI of pineapple. Their effect on yield needs to be determined.Abbreviations ACC oxidase 1-aminocyclopropane-1-carboxylic acid oxidase - CPA 2-(3-chlorophenoxy)propionic acid - AOA aminooxyacetic acid - AVG aminoethoxyvinylglycine - daminozide butanedioic acid mono-(2,2-dimethylhydrazide) - DM dry mass - ethephon [(2-chloroethyl)phosphonic acid] - FM fresh mass - GA gibberellin - EI environmental induction of inflorescence development - IA inflorescence appearance - LSD Fisher's protected least significant difference - MACC malonyl-ACC - NAA naphthaleneacetic acid - PGR plant growth regulator - paclobutrazol (2RS,3RS)-1-(4-chlorophenyl)methyl-4,4-dimethyl-2-(1h-1,2,4-triazol-1-yl)penten-3-ol] - uniconazole (E)-(p-chlorophenyl)-4,4-dimethyl-2-(1,2,4-triazol-1-yl)-1-penten-3-ol - STS silver thiosulfate - M-leaf fourth leaf - Ml-L first leaf younger than M-leaf     

Flower Abscission in Excised Inflorescences of Three Plectranthus Cultivars

Flower abscission induced by ethylene in three Plectranthus cultivars was investigated in order to characterise response to a range of inhibitory and antagonistic compounds. Excised inflorescences were exposed to 100 ml l⁻¹ ethylene gas or placed in various concentrations of ethephon (277, 27.7, 2.77, 0.277 and 0.0277 μM). Flower abscission in Plectranthus was readily induced by applying ethylene gas and by the 277 μM dose of ethephon. Removal of the inflorescences from the ethylene treatment prevented subsequent flower abscission. This implies that ethylene treatment did not induce an autocatalytic production of ethylene. Compounds that are known to compete for the ethylene receptor (100 and 500 ppb 1-methylcyclopropene or 100 and 500 ppm 2,5-norbornadiene) did not reduce abscission in this system. Also, application of the ethylene biosynthesis inhibitor, aminooxyacetic acid at 1 mM, was ineffective at preventing ethylene-induced flower abscission. In contrast, one compound known to block protein production (100 μM cycloheximide) and a non-competitive inhibitor of ethylene action (2 mM silver thiosulfate) did prevent ethylene-induced abscission. We conclude that flower abscission in cut inflorescences of Plectranthus is very likely mediated by endogenous ethylene production, but that control of ethylene-induced flower abscission in this genus can not be readily obtained by most ethylene antagonists that are known to be effective in other systems.     

Effect of substrate-dependent microbial ethylene production on plant growth

Various compounds have been identified as precursors/substrates for the synthesis of ethylene (C₂H₄) in soil. This study was designed to compare the efficiency of four substrates, namely L-methionine (L-MET), 2-keto-4-methylthiobutyric acid (KMBA), 1-aminocyclopropane-1-carboxylic acid (ACC), and calcium carbide (CaC₂), for ethylene biosynthesis in a sandy clay loam soil by gas chromatography. The classic “triple” response in etiolated pea seedling was employed as a bioassay to demonstrate the effect of substrate-dependent microbial production of ethylene on plant growth. Results revealed that an amendment with L-MET, KMBA, ACC (up to 0.10 g/kg soil) and CaC₂ (0.20 g/kg soil) significantly stimulated ethylene biosynthesis in soil. Overall, ACC proved to be the most effective substrate for ethylene production (1434 nmol/kg soil), followed by KMBA, L-MET, and CaC₂ in descending order. Results further revealed that ethylene accumulation in soil from these substrates caused a classic “triple” response in etiolated pea seedlings with different degrees of efficacy. A more obvious classic “triple” response was observed at 0.15, 0.10, and 0.20 g/kg soil of L-MET, KMBA/ACC, and CaC₂, respectively. Similarly, direct exposure of etiolated pea seedlings to commercial ethylene gas also modified the growth pattern in the same way. A significant direct correlation (r = 0.86 to 0.97) between substrate-derived C₂H₄ and the classic triple response in etiolated pea seedlings was observed. This study demonstrated that the presence of substrate(s) in soil may lead to increased ethylene concentration in the air of the soil, which may affect plant growth in a desired direction. Published in Russian in Mikrobiologiya, 2006, Vol. 75, No. 2, pp. 277–283. The text was submitted by the authors in English.     

Respiratory Patterns in Fruit of Pineapple, Ananas comosus, Detached at Different Stages of Development

The postharvest respiratory drifts for six stages of development of pineapple fruit (Ananas comosus cv. Cayenne) ranging from dry flower to senescence are presented. Based upon these data, pineapple is a non-climacteric fruit. Pineapple does produce ethylene gas hut when levels ranging from 0.01 to 1000 μl/l were applied to stage 4 fruits (fruit just at the start of ripening) no respiration or chemical changes were induced which could he interpreted as affecting the ripening processes. A decrease in the oxygen concentration (to 2.5 per cent) resulted in a decrease in the respiration rate. An increased carbon dioxide concentration (up to 10 per cent) had not detectable effect on the respiration rate.     

Role of ethylene and jasmonic acid on rhizome induction and growth in rhubarb (<Emphasis Type="Italic">Rheum rhabarbarum</Emphasis> L.)

Experiments were conducted to elucidate the hormonal induction and regulation of rhizome growth in rhubarb (Rheum rhabarbarum L.). It was found that ethylene is the key regulator of rhizome induction and development. The role of jasmonic acid (JA) and its interaction with ethylene in rhizome induction and growth were also examined. Both ethylene and JA have a significant effect on promoting rhizome formation in vitro. Conversely, the ethylene biosynthesis inhibitor aminoethoxyvinylglycine (AVG) (1.5 μM) inhibited rhizome induction in multiple-shoot clumps in vitro, and suppressed the stimulatory effects of exogenously applied ethephon (1 mg l⁻¹) and JA (10 ng l⁻¹) in promoting mini-rhizome formation, further confirming the role of endogenous ethylene in the process. In addition, rhizome growth was significantly enhanced in the presence of both ethylene and JA (ethephon 1 mg l⁻¹ and JA 10 ng l⁻¹) compared to JA alone. These results suggest that endogenous ethylene is the key regulator of rhizome growth in rhubarb and JA promotes rhizome formation, possibly through inducing endogenous ethylene synthesis.     

Effects of low nitrogen medium on endogenous changes in ethylene, auxins, and cytokinins in in vitro shoot formation from rhizomes of Cymbidium kanran

Summary Shoot formation from rhizome explants of Cymbidium kanran was promoted on Murashige and Skoog (MS) medium: (1) with 1 mgl⁻¹ (4.4μM) 6-benzyladenine (BA) and 0.1 mgl⁻¹ (0.54μM) α-naphthaleneacetic acid (NAA); (2) with ethylene inhibitor (silver nitrate, AgNO₃); or (3) by reducing ammonium nitrate (NH₄NO₃) and potassium nitrate (KNO₃) to 25 and 50%, respectively, of their original concentrations. Shoot formation by BA and NAA was strongly inhibited with the application of ethephon, an ethylene releaser. The ethylene production from the rhizome explants was reduced 30–55% on low nitrogen medium after 1–3 mo. of culture and 52% on BA and NAA medium after 1 mo. of culture compared with explants on standard MS medium. No difference in endogenous auxin (indole-3-acetic acid, IAA) and cytokinin (isopentenyl adenosine, iPA) contents in the rhizomes was found between the treatments. Low ethylene levels were correlated with higher frequency of shoot formation from the rhizomes.     

DEVELOPMENT OF THE INFLORESCENCE AND “CROWN” OF ANANAS COMOSUS AFTER TREATMENT WITH ACETYLENE,NAA, AND ETHEPHON

The vegetative apex of the Masmerah cultivar of pineapple, Ananas comosus (L.) Merr., is dome-shaped and has the typical tunica-corpus organization. Treating the plant with acetylene, NAA or ethephon transforms the vegetative apex to a reproductive one. As early as two days after treatment the apical height and width increase, attaining maximum dimensions in 8–14 days, after which they progressively decrease. During 4–6 weeks, when the full complement of florets has been initiated, the dimensions of the apex are again increased and leaves that form the fruit crown are initiated. A cambium-like zone at the base of the corpus is noticed 2–4 days after the application of acetylene and ethephon. Acetylene and NAA result in a more pointed apex initially, while ethephon results in a more broadened apex. The largest increase in apical height and width is seen when NAA is used. The development of the inflorescence from the time the full complement of florets is formed to fruit maturity is described. Cell multiplication is the main cause of peduncle elongation and inflorescence head enlargement, although towards the later stages of growth, cell enlargement contributes more to the increase in size of the organs.     

Changes in concentrations of soluble carbohydrates during germination of <Emphasis Type="Italic">Amaranthus caudatus</Emphasis> L. seeds in relation to ethylene,gibberellin A<Subscript>3</Subscript> and methyl jasmonate

Unimbibed Amaranthus caudatus seeds were found to contain stachyose, raffinose, verbascose, sucrose, galactinol, myo-inositol, glucose and fructose, while no galactose, maltose and maltotriose was detected. During imbibition, seed concentrations of verbascose, stachyose, raffinose, galactinol, myo-inositol (temporary) and fructose (transient) were observed to decrease; concentrations of galactose and maltose remained fairly constant, while those of sucrose, glucose and maltotriose increased, the increase in sucrose concentration was only temporary. Effects of gibberellin A₃ (GA₃) at 3 × 10⁻⁴ M and ethephon at 3 × 10⁻⁴ M alone or in the presence of methyl jasmonate (Me-JA) at 10⁻³ M on concentrations of soluble sugars during germination of A. caudatus seeds were examined. Me-JA was found to inhibit seed germination and fresh weight of the seeds, but did not affect sucrose, myo-inositol, galactose and maltose concentrations during imbibition for up to 20 h. The exogenously applied GA₃ was observed to enhance germination, stachyose breakdown and glucose concentration after 20 h of incubation. Ethephon stimulated seed germination as well as utilisation of stachyose, galactinol (both after 14 and 20 h) and raffinose (after 14 h of incubation). Although the stimulatory effect of either GA₃ or ethephon on seed germination was blocked by Me-JA; these stimulators increased mobilisation of raffinose and stachyose, but only ethephon enhanced both glucose and fructose after 14 and/or 20 h of incubation in the presence of Me-JA. The maltose concentration was increased by both GA₃ and ethephon alone and in the presence of Me-JA. Of the growth regulators studied, ethephon alone and/or in combination with Me-JA significantly increased the concentrations of glucose, fructose, galactose, maltose and maltotriose. The differences in sugar metabolism appear to be linked to ethylene or GA₃ applied simultaneously with Me-JA.     

Selection of housekeeping genes for gene expression studies on the development of fruit bearing shoots in Chinese jujube (<Emphasis Type="Italic">Ziziphus jujube</Emphasis> Mill.)

Chinese jujube (Ziziphus jujube Mill.) is important economically for its fruit and also bears attractive medical value. Its flower development concurs with the growth of current fruit bearing shoots (FBSs). However, events involved in current FBSs, including FBS differentiation, flower development, and reliable housekeeping gene are still unknown. In this study, the morphology of FBS development and floral formation were examined and expression profiles of several potential internal control genes was determined. These included genes encoding proteins involved in protein translation (translation elongation factor 1α, ZjEF1), protein folding (cyclophilin, ZjCyP) or degradation (ubiquitin extension protein, ZjUBQ), and proteins involved in the structure of the cytoskeleton (β-actin, ZjACT) or nucleosome (histone3, ZjH3). Our results showed that the floral development in early growing FBSs (less than 20 mm in length) or shoot apices was not complete. Among ZjACT, ZjEF1, ZjCyP, ZjUBQ, and ZjH3, ZjH3 was the most suitable housekeeping gene to evaluate FBS development, based on their expression in early growing FBSs, shoot apices, and different organs. These results will be useful for further molecular mechanism study about FBS development in Chinese jujube.     

The role of ethylene and brassinosteroids in the control of sex expression and flower development in <Emphasis Type="Italic">Cucurbita pepo</Emphasis>

In this paper we compare the sensitivity of different squash genotypes to ethylene and brassinosteroids by studying the effects of different ethylene and brassinosteroid treatments on the sexual expression and flower development of different C. pepo genotypes: Bolognese (Bog) and Vegetable Spaghetti (Veg), two contrasting lines for ethylene production and sensitivity, as well as Cora, a standard commercial hybrid. Results have demonstrated that ethylene has a much greater effect on sexual expression and flower development in C. pepo than brassinosteroids. Ethephon increases the number of female flowers per plant and reduces the first male phase of development, while treatments with the ethylene inhibitors AVG and STS reduce the number of female flowers per plant and expand the first male phase of development. The differential response observed between genotypes appears to be related to ethylene production and sensitivity. Bog, which produces more ethylene and is more sensitive to this hormone, responded much better to AVG and STS, reducing the number of female flowers per plant, while Veg, which is characterised by lower production of and sensitivity to ethylene, responded better to ethephon by reducing the first male phase of development and increasing the number of female flowers per plant. The differential abortion of female and male flowers in ethephon, AVG and STS treatments, as well as the occurrence of bisexual flowers in the AVG and STS treated plants of the more ethylene sensitive genotypes, demonstrate that ethylene is also involved in the development of female flowers. Female flower buds require a minimal level of ethylene not only to complete their development and maturation without a premature abortion, but also to arrest the development of stamens in the third whorl and to promote the appropriate growth of the carpels. On the contrary, the role of brassinosteroids in the sexual expression of C. pepo was not so evident. The application of brassinazole, an inhibitor of brassinosteroid biosynthesis slightly changes the production of ethylene in the three analysed genotypes, but those changes have little effect on their sexual phenotypes, and they do not alter the development of the unisexual flowers.     

A genetic map of pineapple (Ananas comosus (L.) Merr.) including SCAR, CAPS, SSR and EST-SSR markers

Despite the paramount importance of pineapple (Ananas comosus L.) in world production and trade of tropical fruits, the genomics of this crop is still lagging behind that of other tropical fruit crops such as banana or papaya. A genetic map of pineapple was constructed using an F2 segregating population obtained from a single selfed F1 plant of a cross A. comosus var. comosus (cv. Rondon, clone BR 50) × A. comosus var. bracteatus (Branco do mato, clone BR 20). Multiple randomly amplified markers (RAPD, ISSR and AFLP) were brought together with SSR and EST-SSR markers identified among sequences uploaded to public databases and with sequence-specific markers (SCAR, SSR and CAPS) derived from random amplified markers. Sixty-three randomly amplified markers (RAPD, ISSR and AFLP) were selected and cloned, resulting in 71 sequences which were used to generate sequence-specific SCAR and CAPS markers. The present map includes 492 DNA markers: 57 RAPD, 22 ISSR, 348 AFLP, 20 SSR, 12 EST-SSR, 25 SCARs, 8 CAPS, and the morphological trait locus “piping”, gathered into 33 linkage groups that integrate markers inherited from both botanical varieties, four linkage groups with markers only from var. comosus and three linkage groups with markers exclusively from var. bracteatus. The relatively higher mapping efficiency of sequence-specific markers derived from randomly amplified markers (50.7%) versus SSR (31.4%) and EST-SSR (28.9%) markers is discussed. Spanning over 80% of the 2,470 cM estimated average length of the genome, the present map constitutes a useful research tool for molecular breeding and genomics projects in pineapple and other Bromeliaceae species.     

<Emphasis Type="Italic">Agrobacterium</Emphasis>-mediated genetic transformation of pineapple var. Queen using a novel encapsulation-based antibiotic selection technique

A protocol for Agrobacterium-mediated transformation of a local ‘elite’ Indian variety (Queen) of pineapple [Ananus comosus (L.) Merr, family Bromeliaceae] has been established using a standard transformation vector (pCAMBIA 1304). High transformation efficiency, expressed as the mean percentage of transgenic micro-shoots regenerated from initial callus explants (20.6%) was achieved using a novel encapsulation-based, antibiotic selection procedure. The Agrobacterium-infected micro-shoots derived from callus explants survived selection in high concentration of hygromycin (60 mg l⁻¹ and beyond) in encapsulated alginate beads. The integration of transgene in hygromycin-resistant shoots and plants was confirmed by histochemical GUS assay, PCR amplification and Southern hybridization. It is possible to eliminate false antibiotic positives in pineapple transformation program to a large extent following this procedure.     

The physiology of ex vitro pineapple (<Emphasis Type="Italic">Ananas comosus</Emphasis> L. Merr. var MD-2) as CAM or C3 is regulated by the environmental conditions

Many plant species grown under in vitro controlled conditions can be used as models for the study of physiological processes. Adult pineapple can display CAM physiology while in vitro it functions as a C3 plant. Ex vitro Ananas comosus has plastic morphology and physiology, both easy to modify from C3 to CAM by changing the environmental conditions. The yield of survival for a rentable propagation protocol of pineapple is closely related with the C3/CAM shift and the associated physiological characteristics. In the present work, ex vitro pineapple plants were divided in two sets and subjected to C3 and CAM-inducing environmental conditions, determined by light intensity and relative humidity, respectively, 40 μmol m⁻² s⁻¹/85% and 260 μmol m⁻² s⁻¹/50%. The results demonstrated that the stress imposed by the environmental conditions switched pineapple plants from C3 to CAM behavior. Comparing to CAM induced, C3-induced pineapple plants showed substandard growth parameters and morphological leaf characteristics but a better rooting process and a higher ABA production, a phenotype closer to adult plants, which are expected to produce fruits in a normal production cycle. We conclude that the upholding of these characteristics is conditioned by low light intensity plus high relative humidity, especially during the first 8 weeks of ex vitro growth. It is expected that the better understanding of pineapple acclimatization will contribute to the design of a protocol to apply as a rentable tool in the pineapple agronomic industry.     

<Emphasis Type="Italic">In vitro</Emphasis> nitrogen nutrition and hormonal pattern in bromeliads

Summary Four cytokinins (CKs), indole-3-acetic acid (IAA) and its ester and amino conjugates, and abscisic acid (ABA) levels of two bromeliads, Ananas comosus (L.) Merril and Vriesea gigantea Gaudich., grown in 5 mM (NH₄)₂SO₄ or urea as the sole nitrogen (N) form, were investigated. In both bromeliads, zeatin (Z) and zeatin riboside ([9R]Z) were the most abundant CKs. In A. comosus, CKs levels decreased drastically (≊ 12 times) after 7 and 30 d in media with ammonium and urea, respectively. After 3 d in media with N, V. gigantea CK levels decreased 30 and 20 times in the presence of ammonium and urea, respectively. N-starved A. comosus and V. gigantea exhibited similar ABA levels, but ABA decreased faster in V. gigantea when plants were transferred to media with N. Free IAA levels decreased until the 15th and 30th day when A. comosus was transferred to a medium with ammonium and urea, respectively. N-starved A. comosus amide, ester, and free IAA amounted to 81%, 14%, and 4%, respectively. There was a transient increase in the proportion of amide IAA and a corresponding decrease of the ester and the free IAA proportion when N-starved plants were transferred to media with N. The relationship between the internal hormonal patterns and the different ecological adaptations of the two bromeliads are discussed.     

Endogenous Auxin is Required but Supraoptimal for Rapid Growth of Rice (Oryza sativa L.) Seminal Roots, and Auxin Inhibition of Rice Seminal Root Growth is Not Caused by Ethylene

The dual effects of auxin and ethylene on rice seminal root growth were investigated in this study. Low concentrations of exogenous indole-3-acetic acid (IAA) had no effect on rice seminal root growth, whereas higher concentrations (≥0.003 μM) were inhibitory. In contrast, low concentrations of the auxin action inhibitor p-chlorophenoxyisobutyric acid (PCIB), ranging from 0.5 to 50 μM, promoted rice seminal root growth, whereas high concentrations of PCIB (≥500 μM) and the polar auxin transport inhibitor 2,3,5-triiodobenzoic acid (TIBA) inhibited rice seminal root growth. These results suggest that endogenous auxin is required but supraoptimal for rapid growth of rice seminal roots. In addition, although rice seminal root growth was inhibited by the exogenous ethylene-releasing compound ethephon or the ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) as well as exogenous IAA, the 50% inhibition of growth (I₅₀) caused by ethephon or ACC was weakened by certain concentrations of the ethylene action inhibitor Ag⁺ (0.016-0.4 μM). However, the I₅₀ caused by exogenous IAA was strengthened by Ag⁺ or the ethylene biosynthetic inhibitor aminoethoxyvinylglycine (AVG) and weakened by certain concentrations of PCIB (0.5-50 μM). Together, the inhibitory mechanisms of auxin and ethylene on rice seminal root growth should be different, and auxin inhibition of rice seminal root growth should not be caused by ethylene. Furthermore, our results indicated that a certain threshold level of ethylene was required to maintain rice seminal root growth, and that ethylene within the threshold may antagonize auxin inhibition of rice seminal root growth.     

An evaluation of the effects of exogenous ethephon,an ethylene releasing compound,on photosynthesis of mustard (<Emphasis Type="Italic">Brassica juncea</Emphasis>) cultivars that differ in photosynthetic capacity

Background  

The stimulatory effect of CO₂ on ethylene evolution in plants is known, but the extent to which ethylene controls photosynthesis is not clear. Studies on the effects of ethylene on CO₂ metabolism have shown conflicting results. Increase or inhibition of photosynthesis by ethylene has been reported. To understand the physiological processes responsible for ethylene-mediated changes in photosynthesis, stomatal and mesophyll effects on photosynthesis and ethylene biosynthesis in response to ethephon treatment in mustard (Brassica juncea) cultivars differing in photosynthetic capacity were studied.     

Expression of <Emphasis Type="Italic">FLOWERING LOCUS T</Emphasis> from <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis> induces precocious flowering in soybean irrespective of maturity group and stem growth habit

The flowering integrator gene FLOWERING LOCUS T (FT) in Arabidopsis thaliana is conserved between diverse plant species and is thought to be the flowering signal ‘‘florigen’’, a universal long-distance mobile signal. In soybean, two FT homologs having a function to induce flowering in Arabidopsis have been identified. In this study, we showed that the expression of FT from Arabidopsis by the Apple latent spherical virus (ALSV) vector promoted precocious flowering and terminated vegetative growth in a wide range of genotypes of soybean, without using a short-day treatment. Four determinate and two indeterminate cultivars, infected with ALSV expressing FT (FT-ALSV), set flower buds on shoot apices and terminated vegetative growth at the fourth- to seventh-node stages under long-day conditions. In contrast, non-infected, healthy plants did not set flower buds on shoot apices at the same stage under the same conditions. After flowering, soybean cultivars infected with FT-ALSV, belonging to different maturity groups and stem growth habits, matured and produced seeds. The results suggest that the basic flowering pathway controlled by FT in A. thaliana might also be conserved in soybean. A system for precocious flowering and shortening of generation time using FT-ALSV would be a useful and novel technology for efficient soybean breeding.     

Effects of ethylene on somatic embryogenesis and galanthamine content in <Emphasis Type="Italic">Leucojum aestivum</Emphasis> L. cultures

The influence of ethylene on in vitro morphogenesis of Leucojum aestivum and galanthamine accumulation was studied. Calli were cultivated on Murashige and Skoog (MS) medium supplemented with 25 μM 4-amino-3,5,6-trichloropicolinic acid (picloram) and 0.5 μM benzyladenine (BA). During incubation under these conditions, callus cultures produced ethylene (9.5 nL/g fresh weight: F.W.) whereas no ethylene was found in somatic embryos cultivated on medium supplemented with 0.5 μM α-naphthalene acetic acid (NAA) and 5 μM zeatin. Application of the precursor of ethylene 1-aminocyclopropane-1-carboxylic acid (ACC) increased ethylene production in both cultures, and decreased callus growth by a factor of 1.2, whereas callus growth was enhanced by a factor of 1.1 in the presence of an inhibitor of ethylene silver nitrate (AgNO₃) or by a factor of 1.2 with an absorbent potassium permanganate (KMnO₄). ACC enhanced the induction of somatic embryos and the development of globular embryos. Removal of ethylene by KMnO₄ during somatic embryogenesis led to the development of plants with greater length. Silver thiosulphate (STS) induced galanthamine production in callus cultures (0.1% dry weight), whereas ACC induced galanthamine production in somatic embryo cultures (2% dry weight).     

Diurnal patterns of carbon dioxide,water vapour,and energy fluxes in pineapple [<Emphasis Type="Italic">Ananas comosus</Emphasis> (L.) Merr. cv. Red Spanish] field using eddy covariance

We analyzed the eddy covariance measurements of momentum, mass, and energy taken daily throughout five consecutive seasonal courses (i.e. 840 d after planting) of a pineapple [Ananas comosus (L.) Merr. cv. Red Spanish] field growing in the Orinoco lowlands. This field provides an opportunity for micrometeorological studies because of the flat and windy site; the seasonal weather including ENSO effects and the Crassulacean Acid Metabolism (CAM) physiology of the crop were additional attributes. Soil CO₂ flux was quantified and added to the net ecosystem exchange in order to obtain the canopy flux (F_C). The canopy CO₂ flux partially followed the four phases of CAM sensu Osmond (1978). The daily pattern of gaseous exchange in pineapple showed a continuum spectrum in which a major proportion of CO₂ uptake occurring during the daytime was common and in which the CAM expression was related to day and nocturnal CO₂ uptake. However, the benefits of CO₂ uptake at low water cost were constrained by the limited nocturnal CO₂ uptake. Seasonal and ontogenetic changes affected the energy exchange as well as the partitioning of available energy into sensible (Q_H) and latent (Q_LE) heat. When the hourly net radiation (Q_Rn) reached its maximum value, latent heat flux (Q_LE) to available energy throughout the vegetative and reproductive stages was 0.65, 0.05, 0.30, 0.11, and 0.33 for the 1997 wet season, 1997/98 dry season, 1998 wet season, 1998/99 dry season, and 1999 wet season, respectively. Throughout the growth period, we found the pivotal role of surface conductance (g _S) in both Q_LE and F_C. Furthermore, the canopy responded to environmental changes. During the wet seasons the g _S was strongly influenced by humidity mole fraction deficit and was usually lower than aerodynamic conductance, whereas during the dry seasons, soil water deficit limited evapotranspiration and production rates. For the fully canopy cover, the hourly trend of marginal water cost of pineapple carbon gain in the dry seasons indicated that g _S became sufficiently efficient to reduce the amount of water transported per unit of carbon gain. In the wet season, the coupling of CO₂ uptake and stomatal conductance was more effective in maintaining a higher proportionality between Q_LE and g _S.     

Plant regeneration by somatic embryogenesis and organogenesis in commercial pineapple (<Emphasis Type="Italic">Ananas comosus</Emphasis> L.)

Summary Axillary and terminal buds from suckers of Ananas comosus cv. Phuket were established on Murashige and Tucker-based (MT) medium with 2.0 mgl⁻¹ (9.8 μM) indolebutyric acid, 2.0 mgl⁻¹ (10.74 μM) naphthaleneacetic acid, and 2.0 mgl⁻¹ (9.29 μM) kinetin, followed by multiplication on Murashige and Skoog-based (MS) medium containing 2.0 mgl⁻¹ (8.87 μM) benzyladenine (BA) to provide a continuous supply of axenic shoots. Leaves, excised from such cultured shoots, produced adventitious shoots from their bases when these explants were cultured on MS medium containing 0.5 mgl⁻¹ (2.26 μM) 2,4-dichlorophenoxyacetic acid (2,4-D) and 2.0 mgl⁻¹ (8.87 μM) BA. Embryogenic callus was produced when leaf explants were cultured on MS medium with 3.0 mgl⁻¹ (12.42 μM) 4-amino-3,5,6-trichloropicolinic acid (picloram). Somatic embryos developed into shoots following transfer of embryogenic tissues to MS medium with 1.0 mgl⁻¹ (4.44 μM) BA. Cell suspensions, initiated by transfer of embryogenic callus to liquid MS medium with 1.0 mgl⁻¹ (4.14 μM) picloram or 1.0 mgl⁻¹ (4.52 μM) 2,4-D, also regenerated shoots by somatic embryogenesis, on transfer of cells to semisolid MS medium with 1.0 mgl⁻¹ (4.44 μM) BA. All regenerated shoots rooted on growth regulator-free MS medium, prior to ex vitro acclimation and transfer to the glasshouse. These studies provide a baseline for propagation, conservation, and genetic manipulation of elite pineapple germplasms.