Production of Flower-Inducing Substance(s) by the Treatment of Lemna Extract with Some Commercial Enzyme Preparations

Flower-inducing substance(s) (assayed with Lemna paucicostata151) was produced by incubation of the centrifuged pellet ofthe homogenate of L. paucicostata 441 (P441) with the commercialenzyme preparations, catalase, cellulase, invertase, lipase,pectinase, peroxidase and proteinase K. These enzyme preparationswere effective even after autoclaving. The flower-inducing activity of the centrifuged supernatantwas also enhanced by incubation with the cellulase preparation,but that of the heat-treated supernatant or pellet was not.Oxygen deprivation (incubation in nitrogen gas) or presenceof ascorbate during the incubation markedly lowered the generationof the flower-inducing activity. These results suggest thatthe flower-in-inducing substance(s) is produced by an oxidativereaction from some heat-stable component in the commercial enzymepreparations by the action of some heat-unstable factor (enzyme?)in the plant. (Received June 6, 1990; Accepted August 16, 1990)     

Production of the Water-Extractable Flower-Inducing Substance(s) in Lemna

Crude water extract of Lemna paucicostata Hegelm., strain 441,had high flower-inducing activity. This activity was heat-stable,but water extract of this plant after heat treatment had verylow activity. The water extract heated immediately after homogenizationalso had low activity, and this activity increased rapidly duringincubation of the plant homogenate before heating even at 0?C.The increase in activity did not occur during separate incubationof the supernatant and pellet obtained by centrifugation ofthe homogenate; some reaction between the components of thesupernatant and pellet may be necessary for production of theheat-stable flower-inducing substance(s). Oxygen deprivation(incubation in nitrogen gas) or presence of ascorbate duringincubation of the plant homogenate markedly lowered the generationof the flower-inducing activity. These results suggest thatthe active substance(s) is produced by an oxidative reaction.No significant difference could be found between photoperiodicallyinduced and non-induced plants in the activity of the waterextract after heat treatment. (Received April 9, 1990; Accepted July 5, 1990)     

Flower-Inducing Activity of Water Extract of Lemna

Flowering of Lemna paucicostata 441 (P441), a sensitive short-dayplant (SDP), was promoted under a near critical photoperiodby the crude water extract of the same plant added to the medium.The extract induced flowering in L. paucicostata 151 (P151),a weakly responsive SDP, under continuous light. The activityfor P151 was greatly promoted by simultaneous application ofbenzyladenine, and the extract of only 0.3 mg fr wt plant addedto 10 ml of assay medium with 1 µM benzyladenine was active.Active substance(s) was similarly obtained from both flower-inducedand non-induced plants, and more or less from all species andstrains of Lemna tested, including P151. However, the extractof short-day strains was more active than that of L. gibba G3(G3), a long-day strain. G3 responded only slightly to the extractof either P441 or G3, whereas P151 responded far more stronglyto the extract of P441 than to that of G3. (Received April 17, 1989; Accepted August 10, 1989)     

Bt毒蛋白在转Bt基因棉中的表达及其在害虫-天敌间的转移

以常规棉泗棉3号作为对照,采用酶联免疫生测法（ELISA）和室内生物测定法,研究了转Bt基因棉新棉33B和GK-12不同组织器官中Cry1Ac或Cry1Ab毒蛋白的表达及其向靶标害虫（棉铃虫）、非靶标害虫（棉蚜）以及天敌（龟纹瓢虫）的传递和影响。研究结果表明,新棉33B各组织器官中Bt毒蛋白的表达量较高,为79.7～1 390.0 ng/g鲜重,GK-012较低为165～2640 ng/g鲜重。在花盛期,新棉33B各组织器官中Bt毒蛋白的表达量依次为：柱头、花 >子房、花瓣>群尖;而5～7叶期的初展嫩叶、现蕾初期的幼蕾及花铃期的幼铃表达量相当,而且与花盛期群尖的表达量没有明显区别。同样处于花盛期的GK-12,其各组织器官中Bt毒蛋白的表达量依次为：花药>柱头>花瓣>群尖>子房;而5～7叶期的初展嫩叶、现蕾初期的幼蕾及花铃期的幼铃表达量相当,而且与花盛期群尖的表达量没有明显差异。常规对照棉的幼铃、花药、柱头以及子房中痕量Bt毒蛋白的存在可能与传粉昆虫等的活动有关。在转Bt基因棉田采集的棉蚜和棉铃虫老龄幼虫,其体内均可检测到Bt毒蛋白;在新棉33B棉田采集的龟纹瓢虫幼虫和成虫体内也可检测出Bt毒素。当以Bt棉田的棉蚜饲喂龟纹瓢虫时,龟纹瓢虫的生长发育、存活以及繁殖等基本没有受到影响。     

A Possible Role for Polyamines in the Repression of Growth of Bradyrhizobium japonicum Bacteroids in Soybean Nodules

Soybean plants (Glycine max L. Merr. cv. Tamahomare) accumulatesufficient putrescine and spermidine in their nodules to inhibitthe growth of bacteroids of Bradyrhizobium japonicum strain138NR. Gas-chromatographic analysis showed that the mature nodulesfrom 35-d-old plants contained approximately 1.5 µmoleseach of putrescine and spermidine per g fresh weight. Water-soluble(free) putrescine and spermidine were present at concentrationsof 0.39 and 0.13 µmoles per g fresh weight, respectively.Cadaverine and spermine were not detected in the nodules. Ina yeast-extract mannitol broth at a pH above 7.0, putrescine,cadaverine, spermidine, and spermine at more than 0.5, 0.2,0.05, and 0.05 mM, respectively, inhibited the growth of thebacteroids. The effect of the polyamines was bactericidal athigher concentrations. More than 95% of bacteroids were notable to form colonies on agar plates that contained 0.5 mM spermidineat pH 7.0. The high sensitivity to polyamines was a unique characteristicof the bacteroidform cells of this strain. The bacteroids losttheir sensitivity to the polyamines within 24 hours after theirisolation from nodules. The cultured cells of this strain multipliedin the presence of 2 mM spermidine or spermine. (Received January 28, 1993; Accepted June 14, 1993)     

Identification of 20-hydroxyecdysone and ecdysone from the pupa of the gypsy moth,Lymantria dispar

Normal and reverse-phase high-performance liquid chromatography in conjunction with radioimmunoassay and mass spectrometry were used to identify the free and conjugated ecdysteroids (after enzymatic hydrolysis) from day-4 pupae of the gypsy moth, Lymantria dispar L. Seven ecdysteroids were searched for, but only 20-hydroxyecdysone (964 ng/g fresh weight) and ecdysone (367 ng/g fresh weight) were detected. Analysis of conjugated ecdysteriods after liberation by hydrolysis also indicated the presence of 20-hydroxyecdysone (21.6 ng/g fresh weight) and ecdysone (2.4 ng/g fresh weight). Neither 26-hydroxyecdysone nor the 3α-epimers of 20-hydroxyecdysone or ecdysone were detected.     

Evidence for Cytokinin in Bacterial Leaf Nodules of Psychotria punctata (Rubiaceae)

Cytokinin activity based on two bioassays was at least 100-fold higher in Psychotria punctata leaf discs with bacterial nodules than in discs without them. Nodulated discs from young leaves yielded 0.4 to 6 μg of cytokinin (zeatin equivalents) per g fresh weight of leaf tissue, whereas non-nodulated discs from the same leaves yielded 0 to 0.003 μg per g fresh weight. These estimates probably include free-base cytokinins and, if present, any nucleoside cytokinins precipitable by acidic silver nitrate. Cytokinin concentrations in Psychotria leaf nodules appear to be higher than normally found in green leaves of other plants. In l-butanol-acetic acid-water (12:3:5, v/v), the one peak of activity chromatographed with an R_F similar to zeatin's, but both number and identity of the active substance(s) remain unknown. These findings suggest that a cytokinin is produced by bacteria in leaf nodules of P. punctata and that it is involved in the symbiosis.     

Morphological Observations and Qualitative and Quantitative Studies of Auxins after Induction of Tobacco Genetic Tumor

When stem segments of tobacco F₁(Nicotiana glaucaxN. langsdorffii)seedlings were cultured on a hormone-free medium in the light,tumorous tissues were induced on the segments within 5 days.The fresh weight of the tissues increased rapidly, with a 10-foldincrease in weight every 6 days. Morphological observationsrevealed the active division of cells and the primordium-likestructures of the teratomas that had been induced during the5 days after cutting of the stem segments. In the light-growntissues of genetic tumors, IAA was revealed to be the predominantauxin by analysis by gas chromatography-mass spectrometry. NormalF₁ seedlings grown in the light contained endogenous IAA ata concentration of 3.4 ng/g fr wt. Five days after cutting,the level of endogenous IAA in stem segments rose to 96 ng/gfr wt or more, and then it decreased to 9.1 ng/g fr wt by 11days. This surge in the endogenous level of IAA may play animportant role in the induction of tobacco genetic tumor. (Received March 30, 1988; Accepted October 31, 1988)     

Nitrate Reduction in Solanum tuberosum L.: Development of Nitrate Reductase Activity in Field-grown Plants

Nitrate reductase activity (in vivo method, substrate non-limiting)in unshaded leaves from the top of the canopy has been determinedfor field-grown potato plants over the course of the growingseason. The pattern of change was almost identical for plantsreceiving no added fertilizer and those receiving 24 g N m^–2.Activity increased to a peak at about 90 days after plantingand declined thereafter. On a fresh weight basis activity wasalways higher in fertilized plants. Nitrate reductase activitywas positively and significantly correlated with leaf proteincontent in high N plants (r² = 0.71; P = 0.05), but poorly correlatedwith both the nitrate content of the leaf lamina and the nitrateconcentration in petiole sap. Up until 90 days after planting(mid-July) there appeared to be a positive relationship betweenincreased activity of nitrate reductase and solar radiation.However, results obtained over two seasons showed that the declinein activity after this time was not consistently linked witha fall in the level of solar radiation. Remobilization of reduced-Nand stored nitrate from leaves and stems accompanied this declinein nitrate reductase activity and in the latter part of theseason appeared to account for all of the N gained by growingtubers. In unfertilized plants nitrate-N accounted for 5 per cent orless of total plant N. Fertilized plants contained up to 25per cent nitrate-N. While nitrate availability limited growthin unfertilized plants, sub-optimal rates of nitrate assimilationin fertilized plants, particularly during the early stages ofpost-emergence growth, may contribute to inefficient use ofacquired nitrate. The carbohydrate status of leaf lamina and petiole sap weremodified by N supply. The soluble sugar and starch contentsof low N leaves were higher than in their high N counterparts.By contrast, the concentration of soluble sugars in petiolesap increased to a higher value in high N samples. Althoughsap sugar levels declined in both treatments towards the endof the season, N application delayed this decline for severalweeks. Solanum tuberosum, nitrate reductase, nitrate assimilation, senescence     

Substance P radioimmunoassay using Nalpha-tyrosyl-substance P and demonstration of the presence of substance P-like immunoreactivities in human blood and porcine tissue extracts.

Sensitive and specific radioimmunoassay for substance P was developed using synthetic substance P and 125I-Nalpha-tyrosyl-substance P. Substance P-human alpha-globulin conjugate was used for production of anti-substance P antisera in rabbits. Synthetic substance P was used as a standard and the dextran-coated charcoal method was employed to separate the free peptide from that bound to antibodies. No cross-reactions by physalaemin and eledoisin observed in this system proved its high specificity to substance P. Nalpha-Tyrosyl-substance P and [Tyr1]-substance P showed the displacement curves indistinguishable from that of the standard substance P. Neither substance P5-11 nor substance P6-11 competed with the tracer at the concentration used. The minimum measurable dose of substance P by the assay system was 2.5-5 pg/incubate. Utilizing the system, human plasma samples from 42 healthy volunteers of both sexes were shown to contain immunoreactive substance P in amounts that averaged 298 pg/ml in male and 251 pg/ml in female. Substance P-like immunoreactivity was also demonstrated in hot-water extracts of porcine duodenum, jejunum, ileum, cecum, middle colon, rectum, pancreas, stomach and pituitary. The highest concentration (379 ng/g wet weight of organ) was found in the pituitary, and the ileum (7.9 ng/g wet weight of organ) and jejunum (1.9 ng/g wet weight of organ) were rich in the contents.     

Feeding‐induced phenol production in Capsicum annuum L. influences Spodoptera litura F. larval growth and physiology

We studied the role of induced plant phenols as a defense response to insect herbivory. Phenolic compounds were induced in Capsicum annuum L., the source of many culinary peppers, after feeding by different stages of the insect pest, Spodoptera litura F. The phenols were identified and quantified using high performance liquid chromatography (HPLC) and effects produced by these phenols on larval development were studied. Vanillic acid was identified in plants challenged by second, fourth, and fifth instar larvae, but not in plants challenged by third instar nor unchallenged plants. Syringic acid production was induced in chili plants infested with second (0.429 ± 0.003 μg/g fresh weight, fourth (0.396 ± 0.01 μg/g fresh weight), and fifth instar (5.5 ± 0.06 μg/g fresh weight) larvae, compared to untreated plants (0.303 ± 0.01 μg/g fresh weight) plants. Leaves surface treated with the rutin deterred oviposition. Dietary exposure to chlorogenic acid, vanillic acid, syringic acid, sinapic acid, and rutin led to enhanced activities of detoxifying enzymes, β‐glucosidase, carboxyl esterase, glutathione S‐transferase, and glutathione reductase in the midgut tissues of all the larval instars, indicating the toxic nature of these compounds. Protein carbonyl content and acetylcholinesterase activity was analyzed to appreciate the role of induced plant phenols in insect protein oxidation and terminating nerve impulses.     

Senescence-Associated Changes during Long-day-induced Leaf Senescence and the Nature of the Graft-transmissible Senescence Substance in Kleinia articulata

Schwabe, W. W. and Kulkarni, V. J. 1987. Senescence-associatedchanges during long-day-induced leaf senescence and the natureof the graft-transmissible senescence substance in Kleinia articulata.— J. exp. Bot. 38: 1741–1755. The long-day-induced senescence in Kleinia articulata leaveswas characterized by a loss in fresh and dry weight, in therate of leaf expansion and progressive loss of chlorophyll inthe detached rooted leaves. Ultrastructural examination of mesophyllcells of leaves from plants grown in continuous light showedthat osmiophilic globules accumulating in the chloroplasts werethe first visible sign of senescence in the organdies. Thesefirst signs of senescence could be detected in very young leavesof plants in continuous light, even before the leaves had expanded.Attempts were made to study the cause of this photoperiodicsenescence which, from previous work, appeared to involve agraft-transmissible substance. Leaves in continuous light showed reduced stomatal opening andextracts from them had very much higher activity in the Commelinastomatal closure assay (ABA-like activity ?) compared with non-senescingleaves grown in short days (8 h). However, even if all the activitywere due to ABA, this on its own does not appear to be the senescencesubstance because a much longer exposure to continuous lightwas required to induce irreversible senescence than to reachmaximum stomatal closure promoting activity in the bioassay.Moreover, severe water stress (high ABA?) did not lead to senescenceunless combined with continuous light or ethylene treatment.It is postulated that while ABA may play an important role inKleinia leaf senescence its lethal effect may not be realizedunless ethylene-induced membrane changes may synergisticallyassist. Key words: Leaf senescence, ABA, Daylength, stomatal movement, Kleinia     

Increased vacuolar and plasma membrane H+-ATPase activities in Salicornia bigelovii Torr. in response to NaCl

The halophyte Salicornia bigelovii Torr. shows optimal growthand Na⁺ accumulation in 200 mM NaCl and reduced growth underlower salinity conditions. The ability to accumulate and compartmentalizeNa⁺ may result, in part, from stimulation of the H⁺ -ATPaseson the plasma membrane (PM-ATPase) and vacuolar membranes (V-ATPase).To determine if these two primary transport systems are involvedin salt tolerance, shoot fresh weight (FW) and activity of thePM- and V-ATPases from shoots in Salicornia grown in 5 and 200mM NaCI were compared. Higher PM-ATPase activity (60%) and FW(60%) were observed in plants grown in 200 mM NaCI and thesestimulations in growth and enzyme activity were specific forNa⁺ and not observed with Na⁺ added in vitro. V-ATPase activitywas significantly stimulated in vivo and in vitro (26% and 46%,respectively) after exposure to 200 mM NaCl, and stimulationwas Na⁺ -specific. Immunoblots indicated that the increasesin activity of the H⁺ -ATPases from plants grown in 200 mM NaCIwas not due to increases in protein expression. These studiessuggest that the H⁺-ATPases in Salicornia are important in salttolerance and provide a biochemical framework for understandingmechanisms of salt tolerance in plants. Key words: Salicornia, H⁺-ATPases, salt tolerance     

Effect of Ammonia on Fruit-body Induction of Coprinus cinereus in Darkness

Coprinus cinereus produced only vegetative mycelia on a potatodextrose medium at 25°C in darkness. When ammonia waterwas added to 5-day-old cultures on the same medium at the concentrationsof 0.05 to 0.4 g NH₃/liter, primordia formed after 3 days inthe dark. Some ammonium salts causing alkalinity in the mediumwere also effective for fruit-body induction, but non-basicammonium salts were not effective unless sodium hydroxide wasadded to make the medium alkaline. The presence of ammonia at least for 24 hr was enough to induceprimordium formation when 5-day-old colonies, after being washed,were treated with ammonia water and then cultured on fresh mediumor even on distilled water. (Received September 10, 1980; Accepted December 29, 1980)     

Involvement of phaseolotoxin in halo blight of beans: transport and conversion to functional toxin

Phaseolotoxin ([N^δ-phosphosulfamyl]ornithylalanylhomoarginine) is produced by Pseudomonas phaseolicola (Burkh.) Dows. in liquid culture. When phaseolotoxin was applied to leaves of bean (Phaseolus vulgaris L.) at 0.1 to 1 nmoles/g fresh weight of leaf by a prick-assay procedure, the characteristic “halo” symptom of bean halo blight disease developed after 24 to 48 hours. At higher concentrations (10-100 nmoles/g fresh weight) the systemic symptoms, which are commonly a feature of diseased plants, also developed after 24 to 48 hours.     

Micronutrient Deficiency Influences Plant Growth and Activities of Superoxide Dismutases in Narrow-leafed Lupins

The effect of copper (Cu), zinc (Zn) or manganese (Mn) deficiencyon the growth and activity of superoxide dismutase (SOD) formswas investigated in seedlings of narrow-leafed lupins (LupinusangustifoliusL.). Plants grown without Zn developed Zn deficiencysymptoms 24 d after sowing (DAS), and those grown without Mnshowed Mn deficiency symptoms 31 DAS. However, plants grownwithout Cu did not show visible leaf symptoms. Shoot dry weightwas decreased by Zn and Mn deficiency 24 DAS, and by Cu deficiency31 DAS. Soluble protein concentration was reduced considerablyby Zn deficiency 24 DAS, but was not affected by Cu deficiencyuntil 31 DAS. In contrast, soluble protein concentration inMn-deficient plants was higher than in control plants 31 DAS.Shoot concentration of micronutrients which were not suppliedto plants decreased significantly, with a simultaneous increasein concentration of one or more of the other nutrients analysed.The activities of total SOD, MnSOD and Cu/ZnSOD on a fresh weightbasis declined drastically in -Cu and -Zn plants 24 DAS. Onthe contrary, the activities of total SOD and Cu/ZnSOD on eithera fresh weight or soluble protein basis increased markedly in-Mn plants 24 DAS, and MnSOD activity increased significantlyin these plants 31 DAS. It was concluded that micronutrientdeficiency (Cu, Zn or Mn) altered the activities of SOD formsdepending on the kind and severity of the deficiency stress.Manipulation of the capacity of plants to tolerate oxidativestress may influence their capacity to tolerate micronutrientdeficiency.Copyright 1999 Annals of Botany Company. Copper,Lupinus angustifolius, manganese, deficiency, superoxide dismutase, zinc.     

ABA Levels and Effects in Chilled and Hardened Phaseolus vulgaris

Leaf abscisic acid (ABA) levels of chilled P. vulgaris weremeasured after 18 h chilling at 5°C, at a saturation deficitof 1.24 g m^–3 (SD), and after chilling in a water-saturatedatmosphere. Changes were also followed during a chill hardeningperiod of 4 d at 12°C, 2.1 g m^–3 SD. It was foundthat hardening resulted in an almost 5. fold increase in ABAlevels after 3 d at 12°C, and this decreased to approximatelycontrol levels on the fourth day. Subsequent chilling of hardenedplants produced no change in ABA levels from that of controlplants (22° C). In contrast, non-hardened plants chilledat 1.24 g m^–3 SD had ABA levels almost 3 times the levelof control plants. However, chilling in a water-saturated atmosphereresulted in a decrease in ABA levels. In addition, the response of leaf diffusion resistance (LDR)to exogenous ABA fed via the transpiration stream was measuredat 5 ° C and 22° C in hardened and non-hardened plants.Use of tritium-labelled ABA was made to calculate the stomatalsensitivity to ABA. It was found that exogenous ABA caused anincreased in LDR at 22°C in both hardened and non-hardenedplants. However, the sensitivity of the hardened plants to ABAwas greater in terms of rate of closure and amount of ABA requiredto close the stomata. At 5°C, however, ABA caused stomatalopening and the maintainance of open stomata in non-hardenedplants. In hardened plants, ABA caused stomatal closure at 5°C.These results are discussed in relation to the locking-openresponse of chilled P. vulgaris stomata. Key words: Chilling, Stomata, ABA, Phaseolus vulgaris     

Cholinergic constituents in plants: Characterization and distribution of acetylcholine and choline

Acetylcholine in plants was identified by gas chromatography/mass spectrometry. Acetylcholine was found in the following species from 13 families: Betula pendula, Codiaeum variegatum, Ilex opaca, Liquidambar styraciflua, Lonicera japonica, Phaseolus aureus, Phaseolus vulgaris, Pisum sativum, Plantago rugelli, Populus grandidentata, Prunus serotina, Rhus copallina, Smilax hispida, Viburnum dilatatum , and Zea mays . Levels of acetylcholine in leaves ranged from a low of 0.14 ± 0.05 (mean ± SEM) nmol (g fresh weight)⁻¹ in I. opaca to a high of 53 ± 6.6 nmol (g fresh weight)⁻¹ in P. aureus . Acetylcholine was found in all tissues examined regardless of the organ (leaves, stems, or roots) or developmental stage (seedlings, mature plants, or seeds). For P. aureus , continuous light exposure increased acetylcholine levels of leaves, and decreased levels in stem when compared to dark controls. Levels of choline, a precursor of acetylcholine, found in leaves ranged from a low of 84 ± 7.0 nmol (g fresh weight)⁻¹ in L. styraciflua to a high of 3700 ± 200 nmol (g fresh weight)⁻¹ in P. aureus . With these findings, three out of the four components of the cholinergic system have now been identified in plants.     

Detection of maize rough dwarf virus by enzyme-linked immunosorbent assay in plant hosts and in the planthopper vector

SVPs were efficiently detected by ELISA in individual male and female insects. Females carried more virus per insect and per unit fresh weight, but no significant difference was detected between males and females in vector efficiency. Of insects positive in ELISA, 15–20% were unable to transmit the virus to host plants. Storage of viruliferous hoppers at-20°C decreased the level of viral antigen detected by half in about 240 days. Subviral particles (SVPs) of maize rough dwarf virus were detected in the planthopper vector Laodelphax striatellus and in maize and barley plants using double antibody sandwich ELISA. In purified preparations diluted in buffer, as little as 36 ng/ml of SVPs was detectable whereas after dilution in extracts of healthy frozen planthoppers the sensitivity was reduced to 50 ng/ml. Freezing the hoppers prior to extraction lowered to one third the background reading due to normal components. Neither the dissociated proteins of the SVP nor the viral double-stranded RNA contributed to the ELISA readings.     

Vegetable Plant Part Relationships. III. Modification of Carrot (Daucus carota L.) Root and Shoot Weights by Gibberellic Acid and Daminozide

Aqueous foliar sprays of N-dimethylaminosuccinamic acid (daminozide)at 2000 p.p.m. and gibberellic acid (GA) at 100 p.p.m. wereapplied 45, 59, 82 and 100 days after sowing to Chantenay carrotswith population densities of 244, 495 and 883 plants m^–2.The plants were harvested on ten approximately weekly occasions;fresh weights were determined and d. wt estimates were obtainedfor the separated shoots (s) and roots (r). Allometric linearregressions of the logarithm of s on that of r at each harvestseparately, clearly showed that GA always increased shoot: rootratio and reduced root yield (by approximately 35 per cent)but could sometimes also increase whole-plant weight. Daminozideincreased root yield (by approximately 7 per cent from 80 tonnesha^–1) and tended to have effects opposite to those ofGA. Daucus carota L., carrot, root weight, shoot weight, N-dimethylaminosuccinamic acid (daminozide), gibberellic acid