De novo differentiation of shoot buds from leaf-callus of Dianthus caryophyllus L. and control of hyperhydricity

A method is described for producing de novo shoots from leaf derived callus of carnation (Dianthus caryophyllus L.). Plants were regenerated in four steps, viz. callus induction, shoot regeneration, removal of hyperhydricity from regenerated shoots and root development. Callus induction medium contained 2,4-D and BAP. Shoot buds were formed when the callus was further subcultured on 2,4-D- and BAP-containing medium, or MS medium without any growth regulators. The shoots so formed were hyperhydric, bushy in appearance with reduced stem length and watery leaves. The normal conformation of shoots was restored by culturing the hyperhydric shoots onto medium supplemented with GA₃ and bactopeptone. The recovered shoots were rooted on MS medium added with NAA (1 mg/l) or IBA (2 mg/l). Regenerated plants with well-developed root and shoot systems were successfully transferred to field conditions after initial acclimation.     

Factors controlling high efficiency adventitious bud formation and plant regeneration from in vitro leaf explants of roses (Rosa hybrida L.)

Studies were conducted to improve adventitious bud regeneration in roses (Rosa hybrida L.), specifically to extend the protocol to different genotypes and to initiate production of multiple shoots per explant. The best results were obtained by using a two-stage procedure where excised leaflets were incubated on Murashige and Skoog (MS) (1962) induction medium with 6.8 μM TDZ plus 0.49 μM IBA in the dark for 7 days and subsequently transferred to an MS-based regeneration medium with 2.22 μM BA plus 0.049 μM IBA exposed to a PPFD of 15 μmol m⁻² s⁻¹ PAR. Bud formation capacity was also significantly affected by the genotype and the environment, such as the use of bottom cooling creating a lower RH in the vessel. The addition of silver nitrate to the induction medium also significantly improved the percentage of regeneration in three genotypes tested. Regenerated shoots failed to elongate when transferred to MS proliferation medium containing 0.5 mg l⁻¹ BA, however maximum bud development and elongation were achieved when kinetin in the range 1–2 mg l⁻¹ was used. Elongated shoots were excised and rooted best on zero growth regulator half-strength MS modified medium. Rooted plantlets were acclimatized under greenhouse conditions for evaluation of somaclonal variation.     

Shoot regeneration via direct organogenesis from in vitro derived leaves of mulberry using thidiazuron and 6-benzylaminopurine

A brief culture of mulberry leaves for 8–10 days on MS medium with 18.17 μM TDZ followed by transfer to 8.88 μM BAP supplemented medium triggered high frequency shoot organogenesis (77.6–89.2%) and favoured shoot elongation in Morus spp. Shoot proliferation was highest in the presence of 2.22 μM BAP with induction of 9.4–10.6 shoots per culture. High frequency of root induction (76.0–86.6%) was observed on medium supplemented with 0.49 μM IBA whereas increase in the level of IBA (4.92 μM) resulted in induction of roots along with development of callus from the base of the shoots. The regenerated plants established in soil at higher frequency in rainy season compared to winter and summer.     

The effect of mycorrhizal symbiosis on the development of micropropagated artichokes

In this work, microrosettes of Cynara cardunculus L. var. scolymus Fiori of the “catanese” type were subcultured in a medium supplemented with 6-benzylaminopurine (BAP) (0.05 mg l⁻¹). For root induction, indoleacetic acid (IAA), α-naphthalene acetic acid (NAA) and indole-3-butyrric acid (IBA) were used at three concentrations: 2, 5 and 10 mg l⁻¹. The highest percentage of rooted shoots was aided by the presence of 10 mg l⁻¹ IAA.Once transplanted in pots, the plantlets were inoculated with 10 g Glomus viscosum strain A6 (AM fungus). Acclimatisation was clearly facilitated by the addition of the AM fungus. Indeed, the mycorrhizal plantlets registered a survival of between 90 and 95% for the rooting shoots and 60% for the non-rooting shoots.The botanical characterization of the material produced was carried out in field and was based on several morphological and productive parameters. Data collected confirm the characteristics of the original cultivar, the efficiency of the in vitro propagation material and the possibility of using this technique in early types of artichoke.     

Micropropagation of Karonda (Carissa carandas) through shoot multiplication

Shoot tips from field grown, mature plants of Carissa carandas cv. Pant Sudarshan were cultured on Murashige and Skoog’s (MS) basal medium supplemented with benzyladenine (BA) and indolebutyric acid (IBA) during different seasons. The maximum sprouting rate was obtained with 1.5 cm long explant collected in spring season (February–March) followed by those collected in summer season (April–June). Shoot proliferation was highest on MS basal media supplemented with 3.0 mg l⁻¹ BA. Rooting of microshoots was noted to be the best in 1/2 MS plus 0.8 mg l⁻¹ IBA and 0.2 mg l⁻¹ naphthalene acetic acid (NAA). The rooted plantlets were successfully acclimatized in vermiculite:sand:soil (1:1:1) potting mixture.     

Shoot induction and plant regeneration from receptacle tissues of Lilium longiflorum

An efficient system has been developed for the in vitro plant regeneration of Lilium longiflorum Thunb. by culturing receptacle sections from flower buds. The sections were cultured on one-half MS medium plus 30 g l⁻¹ sucrose, 8 g l⁻¹ agar, 5.4 μM NAA or 4.9 μM IBA plus 2.2 μM BAP. A section size of 3–4 mm was found to be optimal. After 60 days an average of 41 shoots were formed per explant. More vigorous shoots were obtained by subculturing on hormone-free medium with 20 g l⁻¹ sucrose. Rooting occurred on one-half MS medium with 1.1 μM NAA. Rooted plants were hardened-off in a greenhouse for two months, and normal flowering plants were produced.     

Effect of leaf retention and flavonoids on rooting of Ilex paraguariensis cuttings

The effect of the mature leaf retention and the exogenous application of flavonoids (naringenin, quercetin and rutin at 30 μM for 12 h) was studied for adventitious rooting of Ilex paraguariensis cuttings. Softwood cuttings harvested from young 3-year-old plants and adult 10- and 20-year-old plants were rooted under intermittent fog. A strong correlation (r² = 0.72) between leaf retention and rooting was noted. The highest percentage of adventitious root formation (40%) was obtained when the leaf was artificially removed after 42 days of incubation. This data was supported by the histological analysis which provided anatomical evidence that cuttings have initiated root primordia by 21 days and the regenerated roots emerge through the epidermis after 35 days of incubation. A strong correlation between the position of the leaf and the site of roots regeneration was observed. A 100% of the rooted cutting with a single leaf only formed roots along the leaf axis at the base of the cutting. Quercetin increased the rooting percentage more than three times compared to the control and all flavonoids tested improved the distribution of roots around the stem without impacting the number of regenerated roots per rooted cutting from 20-year-old plants.     

In vitro response of pistachio nodal explants to silver nitrate

The effect of silver nitrate on shoot differentiation and shoot growth was examined in order to improve the regeneration efficiency of pistachio (Pistacia vera L. cv. Kirmizi) in vitro. Nodal explants of in vitro-grown seedlings were used to test various concentrations and combinations of 6-benzyladenine (BA), kinetin (KIN), gibberellic acid (GA₃) and silver nitrate (AgNO₃). Addition of AgNO₃ up to 48.0 μM to the culture medium improved the regeneration frequency and shoot growth, and reduced basal callus formation in all regenerated explants. The highest regeneration frequency (100%) was recorded on Murashige and Skoog (MS) medium containing 9.0 μM BA, 0.2 μM GA₃ and 24.0 or 48.0 μM AgNO₃ in combination. The best proliferation response in terms of both shoot formation and low callus production was obtained in the medium containing a combination of 9.0 μM BA, 0.2 μM GA₃ and 12.0 μM AgNO₃. Regenerated shoots, coming from three cycles of subculturing in proliferation media, were rooted in half-strength MS medium containing 12.0 μM indole-3-butyric acid (IBA). Well rooted plantlets were acclimatized and eventually established in peat and perlite. The development and optimization of an effective micropropagation protocol that is presented in this paper can give an important contribution to improve the quality of pistachio plants and, as a consequence, of orchard production in Middle East countries.     

Micropropagation of the strawberry tree,Arbutus unedo L.

Actively growing shoots of potted greenhouse-grown strawberry tree (Arbutus unedo L.) were initially sterilised and established in basal woody plant medium containing 11.1 μM BA. Optimum shoot proliferation was achieved on a basal WPM containing MS vitamins, sucrose, agar and 22.2 μM BA. Microshoots rooted successfully in basal in vitro medium containing 10 μM IBA or IAA, but their survival rate during acclimatisation was low. Addition of a mixture 1 part peat:4 parts perlite in the basal in vitro rooting medium (1:1 v/v) containing 10 μM IAA resulted in high rooting percentage and plantlets with branched roots. These plantlets were successfully acclimatised. This novel rooting medium can be exploited further due to its potential in commercial applications.     

Protocol for rapid propagation,and to overcome delayed rhizome formation in field established in vitro derived plantlets of Kaempferia galanga L.

Single medium based efficient protocol for rapid propagation, and to overcome the delayed rhizome formation in field established in vitro derived plantlets of Kaempferia galanga L. through in vitro rhizome induction was achieved. MS medium with combination of 8.87 μM N⁶-benzyladenine (BA), and 2.46 μM indole-3-butyric acid (IBA) induced a mean of 6.2 shoots per explant. Addition of 11.7 μM silver nitrate to 8.87 μM BA and 2.46 μM IBA supplemented medium facilitated the highest number of shoots (mean of 8.3 shoots) as well as roots within 60 days. Subculture of isolated shoots on medium with the same concentration of BA, IBA and silver nitrate increased the number to a mean of 12.1 shoots. Silver nitrate enriched medium developed rhizome at the base of shoots. Increase of sucrose concentration (6–8%) in medium with BA, IBA and silver nitrate favoured the best rhizome development. Ninety five per cent of the plantlets survived in field conditions. The plantlets established in field without in vitro developed rhizome (from medium with BA and IBA) did not form rhizome even at 7 months after transplantation. Instead, they developed tuberous roots only. The plantlets with in vitro developed rhizome (on medium having BA, IBA, silver nitrate, and 6–8% sucrose), and that established from conventional way (through splitting of old rhizome) showed no difference in growth of the rhizome. The present study emphasizes the efficacy of silver nitrate and sucrose to develop rhizome in vitro, which enabled to overcome the delayed development of rhizome, and reduced yield of plantlets established in field without in vitro developed rhizome.     

GA4+7 plus benzyladenine reduce foliar chlorosis of Lilium longiflorum

The effectiveness of two commercial formulations of gibberellin (GA) and benzyladenine (BA) for reducing foliar chlorosis on Easter lily (Lilium longiflorum Thunb.) was compared. On a per liter basis, plants were sprayed with 0, 100, 200, or 400 mg (BA equivalent) of Accel (GA₄₊₇:BA of 1:10) or Promalin (GA₄₊₇:BA of 1:1) when the crop leaf area index (LAI)=3. One group of plants was sprayed with 100 mg of Accel or Promalin (BA equivalent) per liter twice: once at LAI=3 and again 3 weeks later. Plants were harvested when the largest flower bud on each plant measured 13 cm in length, stored for 0 or 3 weeks at 2.5°C in the dark, and then moved into a post-harvest evaluation room at 21°C, where foliar chlorosis was monitored for 3 weeks. Senescence of some lower leaves on plants in every treatment was evident at harvest, and incidence of senescence increased during the 21 days of post-harvest evaluation. Cold storage increased the number of leaves senescing during the subsequent evaluation period. Application of Promalin or Accel significantly reduced leaf senescence compared to that of untreated plants. At harvest, 21% of the leaves on untreated plants were senescent, while plants treated with Promalin or Accel averaged 3 or 9% senescent leaves, respectively. Following 7 days of post-harvest evaluation, Promalin was more effective in preventing chlorosis than Accel at the 400 mg l⁻¹ (BA equivalent) level. Following 14 or 21 days of post-harvest evaluation, Promalin was more effective than Accel for the 100 mg l⁻¹ 2× and 400 mg l⁻¹ (BA equivalent) treatments.Plants in all Promalin and Accel treatments were taller than untreated plants 1 week after sprays were applied. At harvest, plants sprayed with Promalin were between 6 and 14 cm taller than untreated plants, but those treated with Accel were the same height as untreated plants.Neither Promalin nor Accel influenced the occurrence of malformed or aborted flowers in this study. However, cold storage significantly increased the number of plants with aborted buds and malformed flowers. Unstored plants averaged 0.16 aborted buds and 0.02 malformed flowers each, while those stored 3 weeks averaged 0.51 aborted buds and 0.18 malformed flowers each.     

Effect of phytohormones on in vitro shoot multiplication and rooting of lime Citrus aurantifolia (Christm.) Swing

Multiple shoots were produced from node explants of lime (Citrus aurantifolia (Christm.) Swing) on MS medium supplemented with 6-benzylaminopurine (BAP), 6-furfurylaminopurine (kinetin) and α-naphthaleneacetic acid (NAA). The highest number of shoots, nine shoots per node, were produced on a medium containing 2 mg l⁻¹ BAP (8.8 μM), 1 mg l⁻¹ kinetin (4.6 μM) and 1 mg l⁻¹ NAA (5.4 μM). Depending on the concentration of BAP and kinetin, NAA either inhibited, stimulated or did not affect shoot multiplication, which also depended on the cytokinin level. Maximum shoot length was obtained from treatments containing 0.5 mg l⁻¹ BAP (2.2 μM) combined with 1 mg l⁻¹ kinetin (4.6 μM) and 0.5 mg l⁻¹ NAA (2.7 μM). The largest leaves of resultant shoots were produced on a medium containing 0.5 mg l⁻¹ each of kinetin (2.3 μM) and NAA (2.7 μM). Transferring in vitro shoots to rooting media containing indole-3-butyric acid (IBA) and NAA produced complete plantlets. The highest rooting percentage was obtained on a medium containing either 1 mg l⁻¹ NAA (5.4 μM) alone or 0.5 mg l⁻¹ NAA (2.7 μM) combined with 2 mg l⁻¹ IBA (9.6 μM), whereas the highest number of roots were produced on a treatment containing both 2 mg l⁻¹ NAA (10.8 μM) and 2 mg l⁻¹ IBA (9.6 μM). Roots elongated most on treatments containing 0.5 mg l⁻¹ of either NAA (2.7 μM) or IBA (2.4 μM). Shoot growth associated with the rooting phase was the highest in response to 2 mg l⁻¹ IBA (9.6 μM) or 0.5 mg l⁻¹ NAA (2.7 μM). Plantlets that survived acclimatization, 82%, exhibited normal growth in soil under greenhouse conditions.     

Double-phase in vitro culture using sorbitol increases shoot proliferation and reduces hyperhydricity in Japanese pear

This study aimed to improve in vitro shoot proliferation efficiency without inducing hyperhydricity in Japanese pear. The shoot number increased at 2.5–10.0 mg l⁻¹ benzyladenine (BA), while shoot fresh mass increased at 1.0 and 2.5 mg l⁻¹ BA. Different macroelement formulation did not affect shoot proliferation, but adding activated charcoal (AC) to the medium inhibited markedly the production of axillary shoots and biomass and many shoots were hyperhydric. Different carbon sources (CS) significantly increased the shoot number and fresh mass, with the best results for shoot proliferation at 20–30 g l⁻¹ sorbitol. With gelling agents, the shoot number increased at 0.4 and 0.6% agar and 0.3% gellan gum, while fresh mass increased at 0.4% agar. The hyperhydric explants were more than 30% at 0.4–0.6% agar and at any concentration of gellan gum. The improved culture (woody plant (WP) supplemented with 20 g l⁻¹ sorbitol, 0.1 mg l⁻¹ 3-indolyl-butyric acid (IBA), 2.5 mg l⁻¹ BA and 0.8% agar) and double-phase culture (the same medium using a double-phase liquid-gelling agent solidified culture system) produced a higher number of axillary shoots than the conventional culture (1/2MS supplemented with 0.1 mg l⁻¹ IBA, 1.0 mg l⁻¹ BA, 30 g l⁻¹ sucrose and 0.8% agar), moreover, double-phase culture had a higher fresh mass than the other cultures.     

Field performance of highbush blueberries (Vaccinium × corymbosum L.) cv. ‘Herbert’ propagated by cuttings and tissue culture

‘Herbert’ highbush blueberry (Vaccinium × corymbosum L.) plants propagated by softwood cuttings (HT) and obtained by micropropagation (TC) of axillary (AX) and adventitious (AD) shoots of 1-year-old in vitro cultures or 11-year-old cultures (SH) were compared. Propagation methods exerted significant influence on nursery and field performance of blueberries. Cutting-derived HT plants grew more slowly, produced significantly less and shorter shoots and were more variable than micropropagated plants. However, the majority of HT plants developed flowers 1 year earlier, flowered more abundantly, bore significantly larger berries than TC plants. There was no clear difference between AX and AD plants. SH-derived plants had smaller berries with the fewest seeds compared to AX and AD-obtained plants. This reveals that culture age had more significant influence than shoot source for the variation observed among micropropagation systems. The present study underlines the necessity of frequent establishment of in vitro cultures of highbush blueberry and carry them out by limited number of passages.     

In vitro production of Prunus necrotic ringspot virus-free begonias through chemo- and thermotherapy

Prunus necrotic ringspot virus (PNRSV)-free Begonia spp. plants were raised from petioles of virus-infected plants using in vitro techniques. The petioles were grown on MS medium supplemented with 0.2 mg/l NAA and 0.2 mg/l BAP (pH 5.8). For rooting, half-strength MS medium without any plant growth regulators was used. On rooting medium, shoots were subjected to chemotherapy (virazole, 2-thiouracil or 6-azauracil) and thermotherapy (38 °C for 16 h light period and 22 °C for 8 h dark period) separately or in combination. Regenerated plants (treated with chemo- and thermotherapy) were indexed for PNRSV by DAS-ELISA and RT-PCR. An amplified product of 785 bp was obtained by RT-PCR in PNRSV-infected plants. Virazole at a concentration of 20 mg/l was found to be more effective (30 and 20% of PNRSV-free plants as indexed by ELISA and RT-PCR, respectively) in comparison to the other chemicals. Thermotherapy for 25 days gave 35 and 25% PNRSV-free plants as indexed by DAS-ELISA and RT-PCR, respectively. A combination of both treatments gave a good number of PNRSV-free plants (67.5 and 57.5% as indexed by DAS-ELISA and RT-PCR, respectively). At higher concentrations all three chemicals were found to be toxic. Thermotherapy for more than 25 days caused browning of leaves and shoots died.     

Rapid in vitro multiplication and conservation of Garcinia indica: A tropical medicinal tree species

A simple and efficient method has been developed for rapid regeneration of plantlets via adventitious bud differentiation on mature seeds of Garcinia indica (Thouars) Choisy, a medicinally important facultative apomictic tropical tree species. High frequency direct shoot proliferation was induced in seed segments cultured on Murashige and Skoog's medium supplemented with cytokinins (BAP, kinetin and TDZ) alone and in combination with auxin (NAA). Amongst the various combinations used, BAP proved to be the most effective. Multiple shoots formed within 4–5 weeks of culture. The shoot forming capacity of the seeds was influenced by the BAP concentration tested (5–50 μM) and optimal response was observed at different concentrations (12.5–50 μM) in different genotypes investigated. Significant differences were recorded in terms of percent response (27.78–100%) as well as average number of shoots per explant (3.49–57.67) among the four genotypes investigated. Elongation of the induced shoots was achieved on MS basal medium containing 0.2% activated charcoal. The induction medium had a profound effect on rate of bud elongation with shoots induced on lower concentrations of BAP showing as much as four-fold elongation within 4 weeks. Proliferating shoot cultures were established by repeatedly subculturing the shoot nodes on MS medium supplemented with 5 μM BAP. Maximum rooting (91.66%) occurred in shoots cultured on half-strength MS medium supplemented with 10 μM IBA. The plantlets were successfully transferred to soil after hardening; the time period from initiation of shoot buds to transplantation being 15–17 weeks. For in vitro conservation, the shoot cultures were maintained on medium supplemented with 0.5 μM BAP and the subculture duration could be enhanced up to maximum of 11 months.     

Factors affecting the shoot multiplication of Persian walnut (Juglans regia L.)

A number of experiments were conducted to identify suitable procedure for in vitro shoot multiplication of Persian walnut (Juglans regia L.). Three different nutrient media (DKW, MS and WPM) and three different gelling agents (Phytagel, Difco Bacto agar and a mixture of Phytagel and Difco Bacto agar) were studied in the first experiment. Driver and Kuniyuki walnut (DKW) medium solidified with 2.2 g l⁻¹ Phytagel was found optimum. Performance of explants was better on DKW medium than on MS and WPM. The DKW and MS media were not significantly different from each other, but both of them were significantly better than WPM, which was a very poor medium for this species. Phytagel alone was significantly better than Difco Bacto agar or Phytagel combined with Difco Bacto agar. In another experiment different concentrations of BA were studied. Medium containing 1.0 mg l⁻¹ BA and 0.01 mg l⁻¹ IBA was the best, although medium containing 0.6 and 0.8 mg l⁻¹ BA were also successful, and utilisation of 0.4 mg l⁻¹ BA and 0.01 mg l⁻¹ IBA was optimum for shoot elongation. Application of different kinds of auxins (IAA, IBA and NAA at 0.01 or 0.1 mg l⁻¹) with 1.0 mg l⁻¹ BA were also studied. Media containing IBA were significantly better than media containing IAA for shoot fresh weight, but neither of them was significantly different from media containing NAA. Application of 0.01 mg l⁻¹ or 0.1 mg l⁻¹ auxin, with 1.0 mg l⁻¹ BA, was not significantly different for shoot multiplication of Persian walnut. The morphology of shoots on media containing 0.01 mg l⁻¹ IBA was the best.     

In vitro plantlet regeneration of Olea europaea ssp. maderensis

Different media were assayed for Olea europaea L. ssp. maderensis Lowe micropropagation. Shoot elongation and propagation was more efficient on DKW medium supplemented with 4.4 μM BA and 0.4 μM IBA. Higher branching was obtained on OM medium supplemented with 18.2 μM zeatin. Rooting was achieved at higher rates on half strength DKW medium supplemented with 20.7 μM IBA. Plants were acclimated to greenhouse and up to now no morphological changes were observed among the regenerated in vitro plants.     

Prohexadione-calcium affects growth and flowering of petunia and impatiens grown under photoselective films

The response of petunia (Petunia x hybrida Vilm.-Andr. ‘Countdown Burgundy’) and impatiens (Impatiens wallerana Hook ‘Accent Orange Tempo’) to Prohexadione-calcium was evaluated under a clear and a far-red light absorbing greenhouse (A_FR) film to investigate the dosage effect of Prohexadione-Ca and to determine if it can overcome the flowering delay under FR deficient greenhouse environments. Prohexadione-Ca reduced stem elongation of petunia and impatiens under A_FR and clear films when applied 3 weeks after germination. Late applications were less effective. In both crops, main stem length decreased in a quadratic pattern as the concentration of Prohexadione-Ca increased. Under both films, 50–100 mg l⁻¹ Prohexadione-Ca resulted in ≈30% shorter petunia plants. Greater concentrations (500 and 1000 mg l⁻¹) resulted in excessively short plants (over 70%). Prohexadione-Ca at 100 mg l⁻¹ delayed anthesis of petunia by 8 and 3 days under the clear film and the A_FR film, respectively during less inductive photoperiods but had no effect during inductive photoperiods. In impatiens, Prohexadione-Ca at 100 mg l⁻¹ delayed anthesis over 10 days under clear or A_FR film. Greater concentrations (200 and 300 mg l⁻¹) inhibited flowering of impatiens. Prohexadione-Ca treatments significantly affected flower color development. Untreated petunia plants had dark burgundy flowers. Prohexadione-Ca treatment increased L*, a*, and C* values and decreased hue angle indicating that the flowers were faded. Flowers of untreated impatiens plants were bright orange color. Prohexadione-Ca at 100 mg l⁻¹ increased L* value and decreased a*, b*, and C* values indicating that significant petal fading had occurred. Flowers of treated plants were nearly white under both films. Although effective in height control, loss of color would be a major limitation to the use of Prohexadione-Ca on flowering crops.     

Analysis of rhythmic growth in holly (Ilex × meserveae) grown in controlled conditions

The influence of 24 h mean air temperature (18.3, 20.6, 23.9 and 25.8 °C) and photosynthetic photon flux (PPF; 0.6, 2.1, 3.7 and 4.7 mol m⁻² d⁻¹) on the growth cycles of vegetative growth in Ilex × meserveae (‘Blue Princess’ S.Y. Hu) was investigated. Plants propagated from top cuttings were grown in greenhouse compartments. The number of unfolded leaves was recorded continuously throughout the experiment. A modified sine function was fitted to collected data and the values for the amplitude and frequency of the growth curves were analysed. The sine function was tested as a method to evaluate the influence of climate on periodically flushing species. Both amplitude and frequency were significantly influenced by air temperature and PPF. The highest frequency of flushing was found at 23.9 °C and 3.7 mol m⁻² d⁻¹. The function resulted generally in a good fit to collected data with R² values above 0.9. Growth curves of all individual plants were categorised with respect to their growth pattern as poor synchronisation within the treatments did not allow analysis of the mean values of the growth curves.