Effects of Ferrous and Phosphate Ions on Flowering in Lemna paucicostata 6746 in Diluted Hutner's Medium

In Lemna paucicostata 6746, P₂ (flower induction period) incontinuous darkness was largely extended by dilution of 1/2H1S medium (Hutner's medium supplemented with 1% sucrose) to1/10 strength. The dilution of 1/2 H1S medium to 1/10 strengthremoved completely the extension of P₁ (pre-flower inductionperiod) and P₂ due to a red light pulse given at the 7th h ofthe dark period, which was observed in 1/2 H1S medium. When iron and phosphate ions were added to 1/10 H1S medium upto the same concentration as in 1/2 H1S medium, the extensionof P₂ was removed completely. The red light pulse-induced extensionof P₁ was observed in 1/10 H1S medium only when iron ions wereadded. It is suggested that iron (Fe^{$ $}) and phosphate ionsare important in determining the rate of photoperiodic flowerinduction in L. paucicostata 6746. (Received December 23, 1983; Accepted June 6, 1984)     

Effect of far-red light pulse on induction and production of flowers in Lemna paucicostata 6746 in darkness

A short-day duckweed, Lemna paucicostata 6746, was exposed tocontinuous darkness at 26^?C, and the changes in the floral parameters(3) due to far-red and/or red light pulse given at various timesof the dark period were studied. Parameters a (vegetative growth rate) and (flowering ratio)were respectively decreased and increased with a far-red lightpulse given at the outset of the dark period. The decreaseda and the increased remained almost unchanged until the 7thhour, but returned to their initial levels thereafter. The far-redlight actions on a and were reversed by subsequent exposureto red light. Parameter P₁ (pre-flower induction period) wasextended by 1 day when far-red and/or red pulse was given atabout the 7th hour of the dark period. A far-jed pulse givenat the outset of the dark period only affected parameter P₂(flower induction period). Although the sensitivity of P₂ tored light increased with time, its sensitivity to far-red lightremained constant and at about the 7th hour was equally sensitiveto far-red and red lights. Both red and far-red pulses givenlater than the 7th hour were increasingly ineffective on P₂.The red/far-red reversibility occurred only for the action onP₂ of the far-red pulse applied during the early dark period.Parameter P₄ (flower production period) varied rhythmicallyin length with a far-red puke, the maximum shortening and extensionbeing induced by the pulse given at about the 7th and 19th hours,respectively. The sensitivity of P₄ to red light also changedrhythmically with an inverse phase angle to the rhythmic responseto farred light, and the far-red and red light actions werereversed respectively by subsequent red and far-red lights. These findings suggested that multiple timing devices includingan hourglass-type clock and a circadian clock are involved induckweed flowering. (Received October 25, 1978; )     

Flower-inducing Activity of Vitamin K in Lemna paucicostata

Vitamins K₁ K₃ and K₅ induced flowering in Lemna paucicostata151, a short-day plant, cultured in 1/10 strength M medium (1/10M medium) under continuous light, and their activity was greatlyintensified by simultaneous application of benzyladenine. Themost active of these was vitamin K₅ L. paucicostata 6746 ismore sensitive to vitamin K₅ than strain 151, but the effectof vitamin K₅ on strain 6746 was not intensified by benzyladenine.The flower-inducing activity of vitamin K₅ was intensified bythe addition of benzoic acid in both strains and by the additionof copper or ferricyanide in Strain 6746, when these chemicalswere added at such low concentrations that they would scarcelyinduce flowering. In strain 6746, vitamin K₅ added to 1/10 M had little effecton flowering under a subcritical photoperiod, while it clearlyinduced flowering under continuous light. In this strain, vitaminK₅ added to full strength M medium, in which this plant wasmore sensitive to short photoperiods than in 1/10 M medium,did not induce flowering even under continuous light, and wasrather inhibitory under short photoperiods. (Received August 14, 1984; Accepted October 16, 1984)     

Effect of red light pulse on induction and production of flowers in a short-day duckweed, Lemna paucicostata 6746, in darkness

Flowering (number of flowers) of a short-day duckweed, Lemnapaucicostata 6746, in continuous darkness at 26^?C was affectedby a red light pulse in various ways depending on the time ofapplication. A conspicuous inhibition and a slight promotionwere respectively caused by the pulse given at the 7th and 19thhours of the dark period. Of the recently introduced floral parameters (4), a (vegetativegrowth rate) and (flowering ratio) were almost unchanged bythe pulse given at any time. P₁ (pre-flower induction period)was extended by one day when the pulse was given at about the7th hour of the dark period. The pulse greatly extended P₂ (flowerinduction period) when given at about the 7th hour of the darkperiod. A pulse given earlier or later was increasingly ineffectiveon P₂. P₄ (flower production period) changed rhythmically (i.e.,was extended or shortened) with the time of the red light pulse,the maximum extension and shortening being induced by the pulsegiven at about the 7th and 19th hours, respectively. Differenttiming mechanisms were suggested as controlling the sensitivitiesto the red light pulse of P₁ and P₂ or P₄. The floral response (number of flowers) vs. the red light pulseapplication time curve was explained in terms of the sum ofthe responses of P₂ and P₄ to the pulse. Floral parameters P₁and P₂ were defined more clearly. (Received September 4, 1978; )     

Effect of temperature on induction and production of flowers in Lemna paucicostata 6746 in uninterrupted and interrupted darkness

Effects of temperatures ranging from 16 to 31^?C on inductionand production of flowers in Lemna paucicostata 6746 were studiedin uninterrupted and interrupted darkness. In uninterrupted darkness, the growth rate (a) and floweringratio () increased and the flower production period (P₄) decreasedas the temperature rose. On the contrary, the pre-flower inductionperiod (P₁) and the flower induction period (P₂) were independentof temperature except that P₂ was remarkably extended at 31^?C.Thus, a, and P₄ may be rate-limited by chemical reactions andP₁ and P₂ by physical reactions. P₂ started at the onset ofdarkness. A red light pulse given 7 hr after the start of the dark periodextended P₁, P₂ and P₄ without modifying a and at any temperature.The pulse extended P₁ by one day irrespective of temperature,and the sensitivity of P₁ to the pulse was constant at all temperatures.The red light pulse caused obvious extensions of P₂ and P₄ at21–26^?C, but no extension at lower and higher temperatures.Thus, the extension of P₁ by red light seems to be rate-limitedby a physical reaction and those of P₂ and P₄ by chemical reactions. (Received September 26, 1978; )     

Spectral Dependence of Night-break Effect on Photoperiodic Floral Induction in Lemna paucicostata 441

A single dark period of longer than 8 hr induced flowering inLemna paucicostata 441 cultured in E medium. Monochromatic lightsof 450, 550, 650 and 750 nm with a half-power bandwidth of 9nm given for 10 min at the 8th hour of a 14-hr dark period inhibitedflowering. The fluence rates required for 50% inhibition were10, 0.5, 0.1 and 3 µmol m^–2. sec^–1, respectively.When applied between the 4th and the 10th hour of the dark period,lights of 450, 550 and 650 nm were inhibitory showing a maximumeffect at the 8th hour. But 750-nm light completely inhibitedflowering when applied at any time during the first 8 hr ofthe dark period. The inhibitory effect of 750-nm light givenat the beginning of the dark period was totally reversed bya subsequent exposure to 650-nm light, and the fluence-responsecurves for the effect of 750-nm light given at the 0, 4th and8th hour were essentially the same. This suggests that the presenceof P_FR is crucial for the floral initiation throughout the first8 hr of the inductive dark period. The role of phytochrome inthe photoperiodic flower induction of L. paucicostata is discussed. (Received January 4, 1982; Accepted March 19, 1982)     

Effects of Exogenous Amino Acids on Iron Uptake in Relation to their Effects on Photoperiodic Flowering in Lemna paucicostata 6746

The flowering of Lemna paucicostata 6746 grown on 14-h photoperiodwas enhanced by the addition of high concentrations of ironto the medium, which also increased the endogenous iron concentration.The addition of asparagine, aspartate, glutamate, -alanine,glycine or serine to the medium also increased the endogenousiron level, resulting in the promotion of flowering. In contrast,the addition of cysteine, cystine, glutamine, arginine, threonineor phenylalanine lowered the endogenous iron level, resultingin the inhibition of flowering. Glycine and asparagine added to the medium during an inductive96-h dark period did not promote iron uptake and had no effecton flowering, but when added during the subsequent 120-h lightperiod, they promoted both iron uptake and flowering response.The increase in the endogenous iron level seems to favor floraldevelopment rather than induction of photoperiodic floweringof Lemna paucicostata 6746. (Received September 8, 1986; Accepted March 31, 1987)     

Effects of some metabolic inhibitors on flowering of Lemna gibba G3, a long-day duckweed

Flowering of Lemna gibba G₃, a long-day duckweed, was inhibitedby adding CuSO₄, AgNO₃, HgCl₂, Na₂WO₄ or iodoacetamide to themedium at the concentrations inducing long-day flowering inLemna paucicostata 6746, a short-day duckweed. This suggeststhat these metabolic inhibitors affected the photoperiodic sensitivityrather than directly affecting flower initiation. Ferricyanidepromoted flowering in both of these short-day and long-day duckweeds. (Received July 7, 1977; )     

Flower-inducing Effect of Benzoic and Salicylic Acids in Various Strains of Lemna paucicostata and L. minor

The flower-inducing activities of benzoic and salicylic acidsadded to the medium differ with the species (Lemna paucicostataand L. minor), and even with the strains used. The type andpH of the medium used, full or 1/10 strength M medium at pH3.8, 4.4 or 5.1, or 1/2 or 1/20 strength NH₄⁺-free Hutner'smedium at pH 5.0, 6.0 or 7.0, also modify their activity. L.paucicostata, strain 151 is the most sensitive of the strainsused to both benzoic and salicylic acids followed by strain381. Such dramatic flowering responses were not obtained withthe other strains, but even strain 321, reportedly insensitiveto benzoic acid, could be induced to flower by adding benzoicacid to a modification of the medium. Benzoic acid is more effectivethan salicylic acid for all strains of L. paucicostata, butthe contrary is true for two L. minor strains tested. A higherpercentage of flowering is obtained in L. paucicostata in 1/2strength NH₄⁺-free Huter'sn medium than in M medium, exceptfor strain 151. When diluted, both media enhance flowering inall L. paucicostata strains. Generally, a lower concentrationof benzoic acid or salicylic acid is enough to induce floweringwhen the pH of the medium is lower. (Received March 30, 1981; Accepted May 16, 1981)     

Flower-Inducing Activity of Lysine in Lemna paucicostata 6746

Flowering of Lemna paucicostata 6746, a typical short-day plant,was induced by culture for 96 or 120 h in nitrogen-free mediumunder continuous illumination. To examine the effects of lysine,we homogenized entire plants of L. paucicostata 151 in a solutionof lysine and the supernatant obtained after centrifugationof the homogenate was added to the medium to give various concentrationsof lysine in the medium. Flowering of strain 6746 in nitrogen-freeor nitrogen-deficient culture medium was effectively promotedby the addition of a lysine-containing supernatant to the medium.The suppressive effect of elastatinal, a protease inhibitor,on the induction of flowering was almost completely reversedby the simultaneous application of a lysine-containing supernatantto the medium. During nitrogen-free culture, the level of endogenousfree lysine, expressed on the basis of the amount of total freeamino acids, increased. Lysine-containing supernatants alsoinduced flowering of plants in nitrogen-rich medium under continuousillumination. These findings suggest that endogenous lysineis involved in the induction of flowering in L. paucicostata6746 on nitrogen-free or nitrogen-deficient medium, as it isin the induction of flowering in L. paucicostata 151 (Received July 29, 1996; Accepted November 18, 1996)     

Flowering Response in Relation to C and N Contents of Pharbitis nil Plants Cultured in Nitrogen-Poor Media

Flowering of seedlings of Pharbitis nil, strains Violet andTendan, cultured in modified White's medium, was promoted bymedium dilution, the critical dark period being shortened byabout 15 min. Dilution of the N source alone was enough to causethe medium-dilution effect. Dilution of the culture medium duringthe day before and on the day of exposure to the dark-period(a total of two days) caused the maximum dilution effect. TheC and N contents of the cotyledons and of the shoot apices changedrapidly in response to medium dilution. In 1/2-strength White'smedium with 1/1,000 strength NO₃^– which was most effectivefor flower promotion, the C-N ratio was highest. In 1/2-strengthmodified White's medium, in which flowering was lowest withthe longest critical dark period, the C-N ratio was lowest.Thus, there is a close relation between flowering response andthe C-N ratio in cotyledons or shoot apices of Pharbitis nil. (Received September 14, 1984; Accepted January 26, 1985)     

Effect of L-Pipecolic Acid on Flowering in Lemna paucicostata and Lemna gibba

L-Pipecolic acid was found to be effective in inducing floweringof Lemna paucicostata 151, 381, 441 and 6746, and of Lemna gibbaG3. When the plants were grown on half-strength Hutner's medium,L-pipecolic acid caused profuse flowering of L. paucicostata151 maintained under 9 and 10 h of light daily. In L. paucicostata441 and 6746, L-pipecolic acid had a strong flower-promotingeffect under a near critical photoperiod. In L. paucicostata381, by contrast, L-pipecolic acid had only a very small effecton flowering. In L. gibba G3 substantial promotion of floweringwas observed under continuous light. When one-twentieth-strengthHutner's medium was used as the basic medium, L-pipecolic acidstimulated flowering in all strains of Lemna examined, evenunder continuous light. When L. paucicostata 151 was grown on one-tenth-strength M mediumor one-twentieth-strength Hutner's medium, the flower-inducingactivity of L-pipecolic acid was greatly enhanced by cytokininunder continuous light. However, when this strain was grownwith 9 h of illumination daily, this synergistic effect of cytokininwas only slight. A short-term (even 1-h) treatment with L-pipecolicacid resulted in flowering, suggesting that L-pipecolic acidis involved in the induction of flowering, rather than its evocation.D-Pipecolic acid also had flower-inducing activity, but itsactivity was 50 times lower than that of the L-isomer. (Received January 23, 1992; Accepted March 9, 1992)     

Inhibition of Flower Induction by Some Inhibitors of Protease in the Short-day Plant Lemna paucicostata 6746 and the Long-day Plant Lemna gibba G3

Four inhibitors of proteases, namely, bestatin, diisopropylfluorophosphate, elastatinal and p-toluenesulfonyl-L-lysinechloromethyl ketone hydrochloride, were examined for their effectson flowering of a short-day plant Lemna paucicostata 6746 anda long-day plant Lemna gibba G3. Each of the inhibitors greatlyinhibited the flowering of Lemna paucicostata 6746 that is normallyinduced by nitrogen deficiency. Bestatin or elastatinal givenonly during the first half of the culture period inhibited theflowering more clearly than when each was given during the latterhalf, suggesting that they inhibited the inductive process(es)involved in flowering rather than development of flower buds.Bestatin or elastatinal greatly inhibited the flowering of Lemnapaucicostata 6746 induced by photoperiodic stimulus, ferricyanideand continuous far-red light. Simultaneous application of thesetwo inhibitors was more effective in the inhibition of photoperiodicallyinduced and ferricyanide-induced flowering than was each inhibitoralone. They also completely inhibited the photoperiodic floweringof Lemna gibba G3. These results suggest that the inductionor activation of some proteases, probably followed by the degradationof some protein(s), is necessary for the induction of floweringin both these plants. (Received November 21, 1989; Accepted February 19, 1990)     

Enhancement of long-day flowering by Mo-deficiency and application of some amino acids and asparagine in the short-day plant Lemna paucicostata 6746

The short-day plant Lemna paucicostata 6746 can be induced toflower on long days (continuous light) by the addition of copper,tungstate or ferricyanide to the medium, and in each case theeffect was greatly enhanced by deleting molybdate from the medium.Treatment with asparagine, aspartate, glutamate, -alanine, glycineorserine, all of which are known to increase the critical daylengthand almost nullify the light-break effect in L. paucicostata6746, enhanced the long-day flowering induced by copper, ferricyanideor Mo-deficiency. (Received October 13, 1978; )     

Flower Induction by Nitrogen Deficiency in Lemna paucicostata 6746

Lemna paucicostata 6746, a short-day plant, produced flowerbuds even under continuous light when cultured in nitrogen-deficientmodified Hoagland medium with 1% sucrose for 3 days or morefollowed by culture on nitrogen-rich medium (either nitrateor ammonium). Flowering was also induced by culture on mediumcontaining 20–100 µM nitrate as the sole nitrogensource for 10 days or more, but not on medium with a low ammoniumconcentration. However, if plants cultured on medium containing5–20 µM ammonium as the sole nitrogen source for10 days were grown in a nitrogen-rich medium for a further 4days, they produced flower buds. Thus, nitrogen deficiency caninduce day length-independent flowering in Lemna paucicoslata6746, but nitrogen is required for the manifestation of flowering. (Received January 31, 1986; Accepted April 24, 1986)     

Absorption of Copper by Lemna as Influenced by Some Factors Which Nullify the Copper Effect on Flowering and Growth

The effect of copper on flowering and growth of Lemna paucicostata6746 and Lemna gibba G3 in a copper-containing medium is nullifiedby the addition of EDTA, ammonium ions or salicylic acid tothe medium or a decrease in its nitrate concentration. Thesefactors were examined for their effects on the absorption ofcopper by the plants. The addition of EDTA to the medium completelyinhibited the absorption of copper in both species, thus eliminatingthe copper effect. Ammonium ions also inhibited copper absorption,their effectiveness rising with their concentration. Loweringthe nitrate concentration in the medium nullified the coppereffect on flowering in L. paucicostata 6746, and the additionof salicylic acid to the medium also nullified the copper effectin L. gibba G3, both without affecting the absorption of copper. (Received June 7, 1982; Accepted August 27, 1982)     

Flower-promoting effect of some amino acids and amides in Lemna paucicostata 6746

Cultures of Lemna paucicostata 6746 were exposed to a single96-hr dark period followed by continuous illumination at 24?1?C.Flowering percentage increased to a maximum 3 days after theend of the dark period and then fell off to 0% on the 5th day.Among 20 amino acids and 2 amides tested, addition of asparagine,aspartate, glutamate, -alanine, glycine and serine clearly increasedthe flowering percentages and retarded the regression of floralbuds by 2–3 days. These substances given after the endof the long dark period were more effective than those givenduring the dark period, suggesting that they favored the flower-producingprocess following the inductive dark process. On the other hand,if the above amino acids or amide were applied under repeatedlight-dark cycles, they shortened the critical dark period by1–2 hr and almost completely nullified the light-breakeffect. They seem to promote the flower-inductive dark process,too. Glutamate, for instance, was effective even at 5 µM, whilethis amino acid is found in the plant body in large quantities.The mechanism of flower promotion by these amino acids and amideremains unknown. (Received June 3, 1976; )     

Ferricyanide Induces Flowering by Suppression of Nitrate Assimilation in Lemna paucicostata 6746

Lemna paucicostata 6746, a short-day plant, produced flowerbuds even under continuous light when cultured for 3 days inferricyanide containing ammonium-free medium followed by cultureon nitrogen-rich medium (either nitrate or ammonium). Dailytreatment with ferricyanide in the absence of ammonium for morethan 8 hours, which completely inhibited nitrate reductase activitywithin 6 hours after the addition to the medium, induced daylength-independentflowering even when the ammonium-rich medium was given duringthe remaining hours. The presence of ammonium for 1 hour atthe middle of the 14-h ferricyanide treatment almost completelysuppressed floral induction. (Received March 6, 1986; Accepted June 3, 1986)     

Growth and flowering of Lemna paucicostata I. General aspects and role of chelating agents in flowering

Lemna paucicostata HEGELM. is normally a short-day plant andflowers only in the presence of a chelating agent (EDTA or EDDHA)in the medium. The plant can be induced to flower even by asingle long night treatment; the flowering percentage, however,increases with further inductive cycles. The length of the criticaldark period depends upon the chelating agent employed in themedium. It is between 10 and 12 hr in the medium containingEDTA and about 8 hr in the EDDHA-supplemented medium. Red lightinterruption in the middle of the dark period—even fora minute—is inhibitory for flowering. Attempts to identify the metal ion(s) chelated reveal that thechelating agents affect flowering by facilitating iron uptake.This is also supported by the fact that the requirement of achelating agent for flowering can be overcome with an excessof iron in the medium. Interestingly, provision of EDDHA andexcess of ferric citrate, together, can bring about floweringeven under long days. ¹Originally HEGELMAIER (1) designated L. paucicostata as a separatespecies; however, THOMPSON (2) and DAUBS (3) have treated itsynonymous to L. perpusilla. More recently, based on physiologicaland chemotaxonomic studies, the distinctiveness of L. paucicostatafrom L. perpusilla has been favoured (4, 5). (Received September 8, 1969; )