Qualitative and semi-quantitative analyses of cytokinins using LC/APCI-MS in combination with ELISA

Application of liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC/APCI-MS) for the analysis of cytokinins was examined. The fragmentation of cytokinins was studied using authentic trans-zeatin (t-Z), trans-zeatin riboside (t-ZR), isopentenyl adenine (i⁶Ade), isopentenyl adenosine (i⁶Ado), benzyl adenine (BAde), benzyl adenosine (BAdo), and kinetin. These cytokinins were effectively ionized by APCI in aqueous acetonitrile. t-Z, i⁶Ade, BAde, and kinetin showed prominent quasi-molecular ions of [M + H]⁺, and ribosylcytokinins clearly showed both [M + H]⁺ and a characteristic fragment ion ([M + H-ribose]⁺), giving some information about their structures. The qualitative and semi-quantitative analyses of cytokinins by LC/APCI-MS were validated in combination with enzyme-linked immunosorbent assay (ELISA) through the analysis of t-ZR in the teratoma of Nicotiana tobacum. t-ZR was conclusively identified and a semi-quantitative estimate of its endogenous levels were provided by the combination of LC/APCI-MS and ELISA. The quantified values obtained by LC/ APCI-MS (single ion detection) and ELISA are in close agreement.     

Identification of Free Cytokinins and the Changes in Endogenous Levels during Tuber Development of Sweet Potato (Ipomoea batatas Lam.)

Trans-zeatin, trans-zeatin riboside and N⁶-(²-isopentenyl)adenosine(i⁶Ado) were found to be the major cytokinins by high performanceliquid chromatographic separation and gas chromatography-massspectrometric analysis in the small tubers of sweet potato (Ipomoeabatatas Lam. cv. Minamiyutaka) with a diameter of about 5 mm.During tuber development cytokinin levels were high in tubershaving a diameter below 12 mm, minimal in tubers with a diameterof 22.5 mm, and then gradually increased as the tuber developed. The role of cytokinins in tuber development of the sweet potatois discussed. (Received May 20, 1982; Accepted August 10, 1983)     

Determination of plant growth substances in liquid endosperm of immature walnut (Juglans nigra) nuts by an ELISA technique

Plant growth substances (PGSs) were analysed in liquid endosperm of black walnut using HPLC and an ELISA procedure. Of all the PGSs studied, we show no GA₃, low levels of cytokinins (io⁶A, i⁶Ade, i⁶Ado) and ABA, and very high level of IAAAbbreviations ABA Abscisic acid: - Ade Adenine: - GA₃ Gibberellic acid: - IAA Indole-3-acetic acid: - i⁶Ade N⁶(²-1) adenine: - i⁶Ado N⁶(²-isopentenyl adenosine: - io⁶A Zeatin riboside:     

Dynamics of production oftrans-zeatin andtrans-zeatin riboside by immobilized cytokinin-autonomous and cytokinin-dependent tobacco cells

Two strains of cultured tobacco cells (Nicotiana tabacum L. cv. Wisconsin 38) differing in their requirement for exogenous cytokinins (cytokinin-dependent and cytokinin-autonomous) were immobilized on polyphenylenoxide (Sorfix) activated with glutaraldehyde. Columns packed with immobilized cells were continually eluted with diluted Murashige and Skoog's medium lacking or supplemented with synthetic cytokinin (6-benzylaminopurine; BA). Purified samples of column eluates were fractionated by HPLC, andtrans-zeatin (t-Z) andtrans-zeatin riboside (t-ZR) content was estimated by enzyme immunoassay. Both cytokinin-autonomous and cytokinin-dependent tobacco cells produced and excretedt-Z and its riboside, and there were significant quantitative differences between the strains. The steady-state excretion rate oft-Z was 19.8 ng · g^–1 dw · h^–1 and 4 ng · g^–1 dw · h^–1, respectively, and that oft-ZR 4 ng · g^–1 dw · h^–1 and 1 ng · g^–1 dw · h^–1, respectively. Exposure of cytokinin-dependent cells to BA after 72 h of starving for this synthetic cytokinin caused temporary increase in excretion of both zeatin and its riboside. After the application of 5 M BA for 24 h, the excretion rate oft-ZR reached 5 ng · g^–1 dw · h^–1 (5-fold increase), and that oft-Z achieved 12 ng · g^–1 dw · h^–1 (3-fold increase). The elevation oft-Z excretion was delayed about 13 h compared witht-ZR excretion, which started increasing almost immediately after BA application. A pulse of BA in lower concentration (1.5 M for 30 h) provoked lower response.     

Reduction of abscisic acid content and induction of sprouting in potato,Solanum tuberosum L., by thidiazuron

The effect of [(N-phenyl-N-1,2,3-thidiazol-5-ylurea)] (thidazuron) on sprouting of potato (Solanum tuberosum L.) tubers and the role of ABA in bud break and subsequent bud growth were studied. Abscisic acid (ABA) was quantitated by enzyme-linked immunosorbent assay (ELISA) from the peel of potato tubers. The ELISA results were also validated by gas chromatography-electron capture detector and confirmed by gas chromatography-mass spectrometry and by a lettuce hypocotyl bioassay. The degree of rest in the tubers was associated with ABA content in the peel. Basal portion (where tuber was attached to mother plant) contained the highest amount of ABA. Thidiazuron reduced ABA content and induced potato tuber sprouting. Exogenously applied ABA stimulated growth of buds that had emerged from dormancy.On leave from the Department of Horticulture, South China Agricultural University, Guangzhou, People's Republic of China.     

The control of bud dormancy in potato tubers. Measurement of the seasonal pattern of changing concentrations of zeatin-cytokinins

A radioimmunoassay, combined with high-performance liquid chromatography, has been used to analyse the zeatin-type cytokinins of potato (Solanum tuberosum L. cv. Majestic) tubers and tuber buds throughout growth and storage. During tuber growth, zeatin riboside was the predominant cytokinin detected in all tissues. Immediately after harvest, the total cytokinin concentration fell dramatically in the storage tissue, largely as a consequence of the disappearance of zeatin riboside. During storage, levels of cytokinins in the storage tissue remained relatively constant, but increased in the tuber buds. In the buds of tubers stored at 2°C there was a 20-to 50-fold increase in total cytokinin over six weeks, coinciding with the natural break of innate dormancy. At 10°C the rise in the level of bud cytokinins was slower, correlating with the longer duration of innate dormancy. Injecting unlabelled cytokinins into tubers in amounts known to induce sprouting gave rise to increases in cytokinin concentrations in the buds of the same order as the increase associated with the natural break of dormancy. Metabolism of injected cytokinins was greater in non-dormant than in dormant tubers. The roles of cytokinin concentration and the sensitivity of the buds to cytokinin in the control of dormancy are discussed.Abbreviations CK cytokinin - FW fresh weight - HPLC high-performance liquid chromatography - RIA radioimmunoassay - tio⁶ade 6-(4-hydroxy-3-methylbut-trans-2-enylamino)-purine=zeatin - tio⁶adeglc⁹ 6-(4-hydroxy-3-methylbut-trans-2-enylamino)-9--D-glucopyranosyl purine=zeatin-9-glucoside - tio⁶ado 6-(4-hydroxy-3-methylbut-trans-2-enylamino)-9--D-ribofuranosyl purine=zeatin riboside - tio⁶ado-[³H]-diol a radioactive derivative of zeatin riboside, synthesised by periodate-oxidation followed by [³H]NaBH₄-reduction - tio⁶AMP 6-(4-hydroxy-3-methylbut-trans-2-enylamino)-9--D-5-phosphoribofuranosyl purine=zeatin riboside 5-monophosphate - t(ioglc⁴)⁶ade 6-(4-O--D-glucopyranosyl-3-methylbut-trans-2-enylamino)-purine=zeatin-O-glucoside     

The control of bud dormancy in potato tubers

Potato (Solanum tuberosum L.) tuber buds normally remain dormant through the growing season until several weeks after harvest. In the cultivar Majestic, this innate dormancy persisted for 9 to 12 weeks in storage at 10° C, but only 3 to 4 weeks when the tubers were stored at 2° C. At certain stages, supplying cytokinins to tubers with innately dormant buds induced sprout growth within 2 d. The growth rate was comparable to that of buds whose innate dormancy had been lost naturally. Cytokinin-treatment did not accelerate the rates of cell division and cell expansion in buds whose innate dormancy had already broken naturally. Gibberellic acid did not induce sprout growth in buds with innate dormancy. We conclude that cytokinins may well be the primary factor in the switch from innate dormancy to the non-dormant state in potato tuber buds, but probably do not control the subsequent sprout growth.Abbreviations tio ⁶ade 6-(4-hydroxy-3-methylbut-trans-2-enyl amino)purine, zeatin - tio⁶ado 6-(4-hydroxy-3-methylbut-trans-2-enyl amino)-9--D-ribofuranosyl purine, zeatin riboside     

Early responses of drought-resistant and -susceptible tomato plants subjected to water stress

Three-week-old seedlings of one drought-susceptible tomato cultivar (Lycopersicon esculentum cv. New Yorker) and two drought-resistant species of tomato (Solanum pennellii andLycopersicon chilense) were subjected to various degrees of PEG 8000-induced water stress from –0.017 to –1.0 MPa for a duration of 24 h so that their early responses to water stress could be compared. Such a comparison would determine if there was a relationship to root cytokinin levels following sudden induction of water stress in the drought-resistant species. Transpiration rates of leaves were monitored throughout the 24-h period, shoots were evaluated for leaf water potential (LWP), and roots were extracted for levels oft-zeatin riboside (t-ZR) and dihydrozeatin riboside (DHZR) using a monoclonal antibody enzyme immunoassay. Transpiration rates were evaluated gravimetrically by difference every 6 h up to 24 h. Transpiration rate decreased with increasing PEG levels and passage of time in all three species, measured at 6 and 12 h, logarithmically in the case of the twoLycopersicon species and linearly in the case ofSolanum. From 12–18 h (while plants were in darkness), transpiration rate was a function of the level of PEG only and not time in all three species. When light resumed from 18–24 h, only 5.pennellii showed no further decrease in transpiration rate over time with increasing PEG. Drought-susceptibleL. esculentum had a stronger linear decrease in LWP with increasing PEG 8000 concentration than the other two species.L. esculentum also had a higher initial transpiration rate than did either of the drought-resistant species. The two drought-resistant species showed less change in LWP with 5.pennellii having a small decrease andL. chilense having little change. OnlyS. pennellii exhibited a decrease in roott-ZR levels, which may imply a role for root cytokinin within the first 24-h exposure to water stress in this species.L. esculentum exhibited no change in roott-ZR. The levels oft-ZR inL. chilense were less than that ofL. esculentum but showed only a slight decrease with increasing PEG.S. pennellii andL. chilense, although both drought-resistant tomato species, showed different patterns of response with respect to pattern of decline in transpiration rate, LWP, and roott-ZR levels.     

Tolerance regions with segmented fermentation processes

Many microbial fermentation processes exhibit different phases (e.g. adaption phase, main growth phase, main production phase). The process variables e.g. the biomass vary randomly about their mean. The experimentalist is interested to know the break points of the different phases, and a tolerance region, i.e. a range of possible values of the process variable that can be considered as normal. This paper deals with statistical methods for determining break points and tolerance regions.List of Symbols a _i intercept in phasei - b _i specific growth rate in phasei - e _t deviation of a measurement in timet - tEX expectation of variableX - r number of phases of fermentation - T _i break point of phaseit - t _ij time of measurementj in phasei - t _n–2.1–/2 quantile oft distribution - Y(t) logarithm of measurement at timet Greek Letters 1 – cover probability of tolerance region - 1 – part covered by the tolerance region - ² variance ofe _t - (·) standard normal distribution - quantile of chisquare distribution     

Genetic manipulation of 6-phosphofructokinase in potato tubers

The aim of this work was to discover whether genetic manipulation of 6-phosphofructokinase [EC 2.7.1.11; PFK(ATP)] influenced the rate of respiration of tuber tissue of Solanum tuberosum L. Transgenic plants were produced that contained the coding sequence of the Escherichia coli pfkA gene linked to a patatin promoter. Expression of this chimaeric gene in tubers resulted in a 14to 21-fold increase in the maximum catalytic activity of PFK(ATP) without affecting the activities of the other glycolytic enzymes. Tubers, and aged disks of tuber tissue, from transformed plants showed no more than a 30% fall in the content of hexose 6-monophosphates; the other intermediates of glycolysis increased threeto eightfold. Fructose-2,6-bisphosphate was barely detectable in aged disks of transformed tubers. The relative rates of ¹⁴CO₂ production from [1-¹⁴C]-and [6-¹⁴C]-glucose supplied to disks of transformed and control tubers were similar. Oxygen uptake and CO₂ production by aged disks of transformed tubers did not differ significantly from those from control tubers. The same was true of CO₂ production, in air, and in nitrogen, for tuber tissue. It is concluded that PFK(ATP) does not dominate the control of respiration in potato tubers.Abbreviations Fru2,6bisP fructose-2,6-bisphosphate - FW freshweight - GUS -glucuronidase - PFK(ATP) 6-phosphofructokinase - PFK(PPi) pyrophosphate: fructose-6-phosphate 1-phosphotransferase     

Cytokinins of sycamore spring sap

Summary The cytokinins present in the spring sap of Acer pseudoplatanus L. were investigated. Ribosyl-trans-zeatin, trans-zeatin and dihydrozeatin were isolated and identified by combined gas chromatography-mass spectrometry (GC-MS). A number of other cytokinin active fractions were observed. One of these was less polar than zeatin and did not behave as any known cytokinin. Two other fractions were more polar than ribosylzeatin and were unstable. A decomposition product of one of these was identified as ribosyl-trans-zeatin by GC-MS. The possible nature of the unstable compounds is discussed. Data on the changes in cytokinin activity of the various fractions during spring 1973 are presented and discussed.Abbreviations GLC gas-liquid chromatography - GG-MS combined gas chromatography-mass spectrometry - KE kinetin equivalents - TLC thin-layer chromatography - TMS trimethylsilyl - tRNA transfer RNA - i⁶ Ade 6-(3-methylbut-2-enylamino)-purine - i⁶ Ado 6-(3-methylbut-2-enylamino)-9--D-ribofuranosyl-purine     

Phytohormone Profiling During Tuber Development of Chinese Yam by Ultra-high performance Liquid Chromatography–Triple Quadrupole Tandem Mass Spectrometry

Changes in agronomic characters and the profile of various endogenous phytohormones during tuber development were studied in Dioscorea opposite (Chinese yam) cv. Guihuai 16. Tuber development exhibited a sigmoidal growth pattern according to the changes in tuber agronomic characters. The growth cycle of yam tuber could be divided into three stages: initiation stage, enlargement stage, and maturation stage. Moreover, the enlargement stage could be separated into three phases—slow growth phase, rapid growth phase, and late growth phase. Endogenous changes in phytohormones were associated with developmental changes in the tubers. The pulses of bioactive gibberellins (such as GA₃ and GA₄) were measured in tubers. The highest contents of GA₃ and GA₄ were reached 90 days after field planting, corresponding to the beginning of the rapid growth phase of tuber enlargement. Changes in trans-zeatin (tZ), jasmonic acid (JA), indole-3-acetic acid (IAA), and abscisic acid (ABA) levels were also observed, and seemed to be related to tuber enlargement at different phases. Continuous increases in JA and tZ contents accompanied tuber enlargement. Transient pulses of both IAA and ABA contents were also observed at the start of tuber rapid growth. Additionally, a second peak level of IAA was detected at the tuber maturation stage. These results suggest GAs play a key role at the beginning of the tuber rapid growth stage, and there is a close relationship between whole tuber enlargement and the contents of JA and tZ. Moreover, it is suggested that IAA and ABA also may be linked to the beginning of tuber rapid growth, and IAA also seems to be correlated to late tuber maturation.     

Investigation on the role of nodule cytokinins in regulation of nitrate reductase activity ofPhaseolus mungo (L.)

The effects of some nodular cytokinis, zeatin (Z), zeatin riboside (ZR), N⁶ (²-isopentenyl) adenine (IPA), and N⁶ (²-isopentenyI) adenosine (IPAS) on nitrate reductase (E.C 1.9.6.1) activity of root nodules ofPhaseolus mungo were investigated. The cytokinis were also tested for their effect on nitrate uptake by nodules. The results show that IPAS is the most effective of all the four cytokinins tested. Z and IPA, which caused higherin vivo activity than ZR and IPAS, stimulated uptake of nitrate by nodules. The other two (ZR and IPAS) while inhibiting uptake showed greaterin vitro activity than Z and IPA. It may be concluded that some cytokinins, in addition to their direct effects on the enzyme, may increase the substrate availability to it, whereas others may have only an direct effect on the enzyme activation or degradation.Deceased.     

Induction of budding on chloronemata and caulonemata of the moss,Physcomitrella patens,using isopentenyladenine

The bud-inducing effect of the cytokinin N⁶-(²-isopentenyl)-adenine (i⁶-Ade) was examined in the moss Physcomitrella patens growing in liquid culture. Under these conditions, buds could be induced on chloronemata as well as on caulonemata. By application of i⁶-Ade, bud-formation was accelerated in both types of tissue. The number of buds, their size and their site of development were dependent on the concentration of the cytokinin in the range of 10^-7 M to 10^-5 M. Moreover, the percentage of caulonema cells increased with a cytokinin concentration of 10^-5 M. These results indicate that chloronema cells may also function as target cells for exogenous cytokinins. The composition of proteins from caulonemata and chloronemata of two different species (P. patens and Funaria hygrometrica), grown on solid medium were compared. No differences could be detected between the protein patterns of caulonemata and chloronemata of the same species while between the two species the differences were obvious.Abbreviations i⁶-Ade N⁶-(²-isopentenyladenine) - Da dalton - SDS-PAGE sodium dodecyl sulfate-polyacrylamide gel electrophoresis     

Pharmacokinetics of anti-ganglioside GD2 mAb 14G2a in a phase I trial in pediatric cancer patients

A phase I trial of a murine anti-ganglioside (GD2) monoclonal antibody (mAb) 14G2a was conducted in 14 neuroblastoma patients and 1 osteosarcoma patient to assess its safety, toxicity and pharmacokinetics in pediatric patients. The pharmacokinetics of mAb 14G2a were biphasic with at ₁ ^2/ of 2.8±2.8 h and at ₁ ^2/ of 18.3±11.8 h. In general,t ₁ ^2/ was dose-dependent with a level of significance ofP=0.036, and it reached a plateau at doses of 250 mg/m² or more. Overall the peak serum levels were dose-dependent atP<0.001. However, they demonstrated an abrupt increase between doses of 100 mg/m² and 250 mg/m². The latter two suggest a saturable mechanism for mAb elimination. In addition, peak serum concentrations were observed earlier at higher mAb doses, which indicates the achievement of a steady state. Thet ₁ ^2/ of mAb 14G2a in children appears to be shorter than in adults. Furthermore, 2 patients demonstrated a considerable decrease int ₁ ^2/ following retreatment with 14G2a. This was paralleled by high human anti-(mouse Ig) antibody levels. This study represents the first comprehensive analysis of murine mAb pharmacokinetics in children and will be useful in the future design of mAb therapy.This work was supported by grants from FDA, FD-R-000377 and NIH U10 CA 28439 and in part by a grant from the general Clinical Research Center program, MOI RR00827, of the National Center for Research Resources, National Institutes of Health. M. M. U.-F. and C.-S. H. were supported in part by a grant from the Children's Cancer Research Foundation, and R. A. R. was supported in part by NIH grant CA 42508     

Partial auxin deprivation affects endogenous cytokinins in an auxin-dependent,cytokinin-independent tobacco cell strain

The dynamics of individual endogenous cytokinins within the growth cycle (subculture interval) of an auxin-dependent and cytokinin-independent cell suspension culture ofNicotiana tabacum L. (strain VBI-0) were determined using high performance liquid chromatography and radioimmunoassay. In cells grown at an optimum auxin concentration the transient maxima of N⁶-(²-isopentenyl)adenine and N⁶-(²-isopentenyl)-adenosine correlated with the onset of cell division. Cultivation of the cells in a partially auxin-deprived medium resulted in ca. tenfold increase of all endogenous cytokinins. A very distinct maximum of N⁶-(²-sopentenyl) adenine appeared at the beginning of subculture. This indicates that a lack of auxin induced an accumulation of cytokinins predominantly in the form of the free bases, which are physiologically more active than the corresponding ribosides.Abbreviations iP N⁶-(²-isopentenyl)adenine - [9R]iP N⁶-(²-isopentenyl)adenosine - t-Z trans-zeatin - t-[9R]Z trans-zeatin riboside - IAA indole-3-acetic acid - NAA 1-naphthaleneacetic acid - 2,4-D 2,4-dichlorophenoxyacetic acid - f.w. fresh weight - SBI subculture interval - C complete medium - PAD partially auxin-deprived medium - RP-HPLC reverse phase high performance liquid chromatography - RIA radioimmunoassay - PAL L-phenylalanine ammonia lyase     

Evidence in support of biosynthesis de novo of free cytokinins

The four cytokinins in the tRNA from Lupinus luteus L. seeds have been purified and identified as ribosyl-cis-zeatin, 2-methylthio-ribosylzeatin, ( ²-isopentenyl)adenosine and 2-methylthio-N⁶-( ²-isopentenyl)adenosine. These structures have been assigned on the basis of their chromatographic mobilities and the spectroscopic data of the parent materials and their silylated derivatives. The tRNA isolated from Populus x robusta Schneid. leaves contained four cytokinins with identical chromatographic properties to those identified in Lupinus luteus seed tRNA. No evidence was obtained for the presence, in tRNA, of the naturally occurring free cytokinins identified in these plant species, dihydrozeatin (Lupinus luteus) and N⁶-(2-hydroxybenzyl)adenosine (Populus x robusta). This is evidence in support of the possibility that free cytokinins can arise by biosynthesis de novo and are not exclusively by-products released intact during tRNA turnover.     

Regeneration of resistance and ion transport in rabbit corneal epithelium after induced surface cell exfoliation

Summary Exposure of the rabbit corneal surface to a 20-m digitonin-0.9% NaCl solution leads to permeabilization of the most superficial cells of the stratified epithelium. The devitalized cells exfoliate spontaneously from the corneal surface. Detergent exposure limited to 4–8 min leads to permeabilization and rapid exfoliation of a monolayer of surface cells. Consistent with the presence of the epithelial paracellular permeability barrier in this cell layer, their permeabilization results in complete loss of transepithelial resistance (R _t ). Within minutes after detergent removal an initial recovery ofR _t can be noticed indicating generation of a new paracellular permeability barrier by the viable subsurface cells. This recovery proceeds rapidly andR _t reaches within 70 min a maximum equal to > 90% of the preexfoliation values (=2.43 k·cm²,n=22). TheR _t recovery is fully blocked in a reversible manner by 10 m dihydrocytochalasin B. The recovery is not affected by inhibition of protein synthesis with 5 m cycloheximide. When the ocular surface is treated again with digitonin the permeabilization and exfoliation of a monolayer of cells and loss ofR _t are repeated. After the second detergent exposure an initial recovery ofR _t occurs as before within minutes. However, the pace ofR _t recovery is much slower: 4–5 hr are required to reach a stable maximalR _t values amounting to about 73% of initial control. This recovery can be fully blocked by 5 m cycloheximide indicating that protein synthesis is required for generation of tight junctions by the second subcellular layer. With only a fraction ofR _t recovered, short-circuit currents amounting to, at least, 50% of control values and attributable in part to cell-to-tear movement of Cl^– through the apical surface can be measured. This suggests that apical-type Cl^– channels are either present in the apically facing membrane of subsurface cells or that they are rapidly inserted in it from preexisting intracellular pools immediately following the devitalization of the surface cells by digitonin.     

Analysis of Phytohormone Profiles during Male and Female Cone Initiation and Early Differentiation in Long-shoot Buds of Lodgepole Pine

In lodgepole pine (Pinus contorta Dougl. ex Loud. var. latifolia Engelm), cone initiation and gender differentiation are site-specific in long-shoot buds, with female cones in the distal portion and male cones in the proximal portion. By using high-performance liquid chromatography–electrospray ionization tandem mass spectrometry (HPLC–ESI–MS/MS) in multiple-reaction monitoring (MRM) mode, cytokinins, indole-3-acetic acid (IAA), abscisic acid (ABA), and their selected metabolites were investigated in developing long-shoot buds from multiple genotypes. Spatially, higher concentrations of trans-zeatin riboside (t-ZR) and dihydrozeatin riboside (dhZR) existed in the distal parts of long-shoot buds, whereas concentrations of isopentenyl adenosine (iPA), IAA, ABA glucose ester (ABA-GE), and phaseic acid (PA) were higher in the proximal parts in all investigated genotypes. In long-shoot buds of genotypes with a history of high female cone yield, concentrations of t-ZR and the ratio of zeatin-type to isopentenyl-type cytokinins were higher in the entire buds, whereas dhZR or IAA was higher in either the distal or the proximal part, respectively. In low female cone yielding genotypes, concentrations of c-ZR, iPA, ABA-GE, and PA were higher in both of the parts. Temporally, concentrations of several hormone-related compounds showed obvious changes in late June and late July, prior to male and female cone bud differentiation. This study reveals that the local hormonal status in a long-shoot bud at specific developmental stages may play an important role in gender determination and cone yield.     

Early responses of drought-resistant and -susceptible tomato plants subjected to water stress

Three-week-old seedlings of one drought-susceptible tomato cultivar (Lycopersicon esculentum cv. “New Yorker”) and two drought-resistant species of tomato (Solanum pennellii andLycopersicon chilense) were subjected to various degrees of PEG 8000-induced water stress from ?0.017 to ?1.0 MPa for a duration of 24 h so that their early responses to water stress could be compared. Such a comparison would determine if there was a relationship to root cytokinin levels following sudden induction of water stress in the drought-resistant species. Transpiration rates of leaves were monitored throughout the 24-h period, shoots were evaluated for leaf water potential (LWP), and roots were extracted for levels oft-zeatin riboside (t-ZR) and dihydrozeatin riboside (DHZR) using a monoclonal antibody enzyme immunoassay. Transpiration rates were evaluated gravimetrically by difference every 6 h up to 24 h. Transpiration rate decreased with increasing PEG levels and passage of time in all three species, measured at 6 and 12 h, logarithmically in the case of the twoLycopersicon species and linearly in the case ofSolanum. From 12–18 h (while plants were in darkness), transpiration rate was a function of the level of PEG only and not time in all three species. When light resumed from 18–24 h, only 5.pennellii showed no further decrease in transpiration rate over time with increasing PEG. Drought-susceptibleL. esculentum had a stronger linear decrease in LWP with increasing PEG 8000 concentration than the other two species.L. esculentum also had a higher initial transpiration rate than did either of the drought-resistant species. The two drought-resistant species showed less change in LWP with 5.pennellii having a small decrease andL. chilense having little change. OnlyS. pennellii exhibited a decrease in roott-ZR levels, which may imply a role for root cytokinin within the first 24-h exposure to water stress in this species.L. esculentum exhibited no change in roott-ZR. The levels oft-ZR inL. chilense were less than that ofL. esculentum but showed only a slight decrease with increasing PEG.S. pennellii andL. chilense, although both drought-resistant tomato species, showed different patterns of response with respect to pattern of decline in transpiration rate, LWP, and roott-ZR levels.