Photoautotrophic growth response of in vitro cultured coffee plantlets to ventilation methods and photosynthetic photon fluxes under carbon dioxide enriched condition

Effects of two ventilation methods (forced and natural) and two photosynthetic photon fluxes (PPF, 150 and 250 μmol m⁻² s⁻¹) on the photoautotrophic growth of in vitro cultured coffee (Coffea arabusta) plantlets were investigated. Number of air exchanges was 2.7, 5.9 and 3.9 h⁻¹ for forced low rate, forced high rate and natural ventilation, respectively. Single node cuttings of in vitro cultured coffee plantlets were cultured on Florialite, a mixture of vermiculite and cellulose fibers with high air porosity, emerged in liquid half strength basal MS medium, without sucrose, vitamins and plant growth regulators. The study included 40 days in the in vitro stage and 10 days in the ex vitro stage. Mean fresh and dry weights, leaf area, shoot and root lengths and net photosynthetic rate per plantlet were significantly greater in forced high rate treatments compared with those in natural and forced low rate treatments. PPF had a distinct effect on shoot length suppression and root elongation of coffee plantlets in forced high rate treatments. The control of carbon dioxide concentration inside the culture box according to the plant demand when growing was easy with the forced ventilation method in photoautotrophic micropropagation. This revised version was published online in June 2006 with corrections to the Cover Date.     

Photoautotrophic growth of sugarcane plantlets <Emphasis Type="Italic">in vitro</Emphasis> as affected by photosynthetic photon flux and vessel air exchanges

Summary Explants of sugarcane, a C₄ plant, were cultured in vitro for 18d on Floridalite (a solid cube consisting of vermiculite and cellulose fibers) used as supporting material with sugar-free Murashige and Skoog liquid medium with double-strength KH₂PO₄, MgSO₄, FeSO₄, and Na₂-EDTA in the vessel with enhanced natural ventilation. CO₂ concentration in the culture room was kept at 1500 μmol mol⁻¹ (four times the atmospheric CO₂ concentration) during the photoperiod. A factorial experiment was designed with two levels of photosynthetic photon flux (PPF) and three levels of N (number of air exchanges of the vessel). The results were compared with those in the control treatment (photomixotrophic culture using sugar-containing agar medium under low PPF and low N). PPF and N showed significant positive effects on the growth of sugarcane plantlets in vitro. In the photoautotrophic (using sugar-free medium) treatments with relatively high PPF (200–400 μmol m⁻² s⁻¹) and high N (2–10 h⁻¹), the growth of plantlets was four to seven times greater than that in the control. Also, the culture period for multiplication and rooting was shortened from 30 d in the control to 18 d or less in the photoautotrophic, high PPF, and high N treatments. Use of porous supporting material in photoautotrophic treatments promoted rooting and plantlet growth significantly.     

Effects of sucrose concentration,supporting material and number of air exchanges of the vessel on the growth of in vitro coffee plantlets

Growth of coffee (Coffea arabusta) plantlets cultured in vitroas affected by sugar, types of supporting material and number of air exchanges of the vessel was investigated. Single node cuttings of in vitro coffee plantlets were cultured on half strength MS medium with or without 20 g l⁻¹ sucrose. Two types of supporting material, agar and Florialite, and two levels of air exchange expressed by number of air exchanges per vessel, 0.2 and 2.3 h⁻¹, were studied. At the end of a 40-day culture period, fresh weight, shoot length, root length and leaf area of plantlets when cultured on Florialite soaked in sugar-free medium and under the higher number of air exchanges were greater than those in sugar containing medium. Callus was observed at the shoot base of plantlets grown on agar medium containing sucrose. Photosynthetic ability of coffee plantlets in vitro was also significantly increased when grown on sugar-free medium with the high number of air exchanges and Florialite as a supporting material. This revised version was published online in June 2006 with corrections to the Cover Date.     

In vitro and ex vitro growth of grapevine rootstock `5BB' as influenced by number of air exchanges and the presence or absence of sucrose in culture media

Single node cuttings of grapevine rootstock `5BB' were cultured for 5 weeks under different numbers of air exchange (0, 0.8, 1.5, 2.5 and 4.4 h ^–1) with or without sucrose in the medium. To determine the effect of number of air exchanges related to presence or absence of sucrose, in vitro growth, net photosynthetic rate, stomata characteristics and ex vitro growth were investigated. Increasing numbers of air exchanges accelerated plantlet growth regardless of the presence or absence of sucrose in the medium; under the same number of air exchanges, plantlet growth and net photosynthesis were greater in sucrose-containing medium compared with sucrose-free medium. However, a high number of air exchanges (4.4 h^–1) inhibited the growth and photosynthesis in sucrose-containing medium compared to sucrose-free medium. A higher number of stomata was observed in sucrose-containing medium, and larger size stomata was observed in sucrose-free medium. These results emphasize the importance of ventilation (increased number of air exchanges) during in vitro culture, for ex vitro plantlet survival was greatly affected by with or without air exchanges. Without air exchanges ex vitro plantlet survival was less than 70%, while air exchanges increased ex vitro survival by more than 90% with greater growth.     

High photosynthetic photon flux and high CO2 concentration under increased number of air exchanges promote growth and photosynthesis of four kinds of orchid plantlets in vitro

Summary In vitro plantlets of Phalaenopsis ‘Happy Valentine’, Neofinetia falcate Hu, Cymbidium kanran Makino, and Cymbidium goeringii Reichb. f. were grown under photoautotrophic [high photosynthetic photon flux (PPF), high CO₂ concentration, and increased number of air exchanges] and heterotrophic (low PPF, low CO₂ concentration, no air exchanges) culture conditions. After 40 d of culture, a significant difference in plantlet growth was observed between the two cultures. Total fresh and dry mass were on average 1.5 times greater in photoautotrophic culture than in heterotrophic culture. Higher net photosynthetic rates were also observed for Phalaenopsis in photoautotrophic culture. In photoautotrophic culture, little difference was observed in air temperature between the inside and outside of the culture vessel, whereas in heterotrophic culture, air temperature inside the culture vessel was 1–2°C higher than that outside the culture vessel. Relative humidity inside the culture vessel was remarkably different between the two cultures: 83–85% in photoautotrophic culture and 97–99% in heterotrophic culture. These results indicated that growth and net photosynthetic rate of in vitro orchid plantlets were susceptible to the culture environments such as PPF, CO₂ concentration, relative humidity (RH), and the number of air exchanges, which would allow a more efficient micropropagation system for these orchid plants.     

Exogenous sucrose can decrease <Emphasis Type="Italic">in vitro</Emphasis> photosynthesis but improve field survival and growth of coconut (<Emphasis Type="Italic">Cocos nucifera</Emphasis> L.) <Emphasis Type="Italic">in vitro</Emphasis> plantlets

Summary Coconut (Cocos nucifera L.) plantlets grown in vitro often grow slowly when transferred to the field possibly, due to a limited photosynthetic capacity of in vitro-cultured plantlets, apparently caused by the sucrose added to growth medium causing negative feedback for photosynthesis. In this paper, we tested the hypothesis that high exogenous sucrose will decrease ribulose 1,5-bisphosphate carboxylase (Rubisco) activity and photosynthesis resulting in limited ex vitro growth. Plantlets grown with high exogenous sucrose (90 gl⁻¹) had reduced photosynthetic activity that resulted in a poor photosynthetic response to high levels of light and CO₂. These plantlets also had low amounts of Rubisco protein, low Rubisco activity, and reduced growth despite showing high survival when transferred to the field. Decreasing the medium’s sucrose concentration from 90 to 22.5 gl⁻¹ or 0 gl⁻¹ resulted in increased photosynthetic response to light and CO₂ along with increased Rubisco and phosphoenolpyruvate carboxylase (PEPC) activities and proteins. However, plantlets grown in vitro without exogenous sucrose died when transferred ex vitro, whereas those grown with intermediate exogenous sucrose showed intermediate photosynthetic response, high survival, fast growth, and ex vitro photosynthesis. Thus, exogenous sucrose at moderate concentration decreased photosynthesis but increased survival, suggesting that both in vitro photosynthesis and exogenous sucrose reserves contribute to field establisment and growth of coconut plantlets cultured in vitro.     

Growth of In vitro banana (Musa SPP.) shoots under photomixotrophic and photoautotrophic conditions

Summary In vitro banana (Musa spp.) shoots were cultured under photomixotrophic (30 gl⁻¹ sucrose and 0.2 h⁻¹ number of air exchanges of culture vessels) and photoautotrophic (0 gl⁻¹ sucrose and 3.9 h⁻¹ number of air exchanges) conditions for 28 d in 370 cm³ Magenta boxes (GA7-type) containing 70 ml of half-strength Murashige and Skoog (MS) medium with 22.2 μM N⁶-benzyladenine (BA). The effects of varying CO₂ concentration (475 or 1340 μmol mol⁻¹) and light intensity (photosynthetic photon flux (PPF) of 100 or 200 μmol m⁻² s⁻¹) were investigated. Fresh and dry weights of banana shoots grown photomixotrophically were significantly greater on day 28 than those grown photoautotrophically. Photoautorophic shoots had a larger number of unfolded leaves and greater leaf area than photomixotrophic plants by days 14 and 28, regardless of CO₂ concentration. The shoot fresh and dry weights on day 14 in photoautotrophic conditions were significantly greater at PPF of 200 μmol m⁻² s⁻¹ than at 100 μmol m⁻² s⁻¹. The increase in net photosynthetic rate of photoautotrophic banana shoots was significant compared with photomixotrophic shoots. The multiplication ratio of in vitro banana shoots grown photoautotrophically in a 28-d culture period was the greatest at 100 μmol m⁻² s⁻¹ PPF and 475 μmol mol⁻¹ CO₂.     

Optimizing environmental factors for large-scale multiplication of chrysanthemum (<Emphasis Type="Italic">Chrysanthemum grandiflorum</Emphasis>) in balloon-type bioreactor culture

Summary We have developed optimum culture conditions for the large-scale propagation of chrysanthemum in balloon-type bioreactors to achieve vigorous growth and quality. The effects of NH ₄ ⁺ /NO ₃ ⁻ ratio, air volume, air temperature, photosynthetic photo flux, and an inoculation density on the growth and quality of plantlets were investigated. The best production conditions were an NH ₄ ⁺ :NO ₃ ⁻ ratio of 20∶40 mM, air exchange of 0.1 vvm min⁻¹, air temperature 25°C, photosynthetic photo flux (PPF) at 100 μmol m⁻² s⁻¹, and an inoculation density of 40 nodes Chrysanthemum grandiflorum. Under each of these conditions, the maximum growth rate reached 279.0, 260,0, 20.0, 23.3, and 94.5 (g-fresh weight per plantlet d⁻¹), respectively, at 12 wk of culture. These results specify the key environmental factors that can be regulated to improve the quality and quantity of flowers and increase yield in large-scale bioreactor cultures of chrysanthemum.     

Leaf ultrastructure, photosynthetic rate and growth of myrtle plantlets under different in vitro culture conditions

The in vitro rooting of myrtle (Myrtus communis L.) plantlets was performed in containers with gas permeable (V) and non-permeable (C) closures characterized by a different number of gas exchanges (1.4 and 0.3 h^{− 1}, respectively). The rooting was induced on Perlite, soaked with half strength Murashige and Skoog (MS) medium with 0.5 mg dm⁻³ IBA, either with and without 15 g dm⁻³ of sucrose. During the rooting phase, it was demonstrated that C cultures without sucrose (C−) negatively affect the growth of myrtle plantlets. The net photosynthetic rate and the starch content showed the lowest values in C cultures with and without sucrose (C+ and C−) while chlorophyll a content did not vary among treatments, therefore it could not be considered an indicative parameter to evaluate the autotrophic metabolism in myrtle plantlets. Electron microscopy and image analysis were employed to evaluate the leaf ultrastructure at three sample dates. Plantlet rooted in vented vessels with and without sucrose (V+ and V−) showed chloroplasts with numerous starch inclusions, while several osmiophilic plastoglobules (frequently related with leaf senescence) were found in chloroplast of leaf cells of C− myrtle plantlets.     

Growth, morphogenesis, and essential oil production in Mentha spicata L. plantlets in vitro

To date, plantlet culture has not been explored as a means to obtain secondary metabolites in vitro. However, plantlets readily produce desirable secondary metabolites, which may not be produced in cell suspension or callus cultures. To optimize plantlet growth in vitro, the influences of various physical environments on the growth (fresh weight), morphogenesis (leaf, root, and shoot number), and volatile carbon metabolites (i.e. monoterpene, (−)-carvone) of Mentha spicata L. (spearmint) plants were studied. The carvone content in different portions of sterile plantlets was analyzed. Carvone was only produced from the foliar regions of cultured plantlets and was absent in the callus and roots. The influence of physical support (e.g., agar, glass gravel, liquid, platform or sponge), frequency of media replacement, and culture vessel capacity on spearmint plantlets growth and carvone production was tested. A comparative study was conducted testing the growth, morphogenesis, and secondary metabolism occurring with three different spearmint cultivars grown in either culture tubes containing 25 ml agar medium or in an automated plant culture system (APCS; a sterile hydroponics system) employing a 1-l medium reservoir. Increasing the number of media immersions (4, 8, 12 or 16 immersions d⁻¹) of plantlets growing in the APCS increased growth and morphogenesis responses. Generally, higher culture growth rates resulted in lower carvone treatment⁻¹ (mg carvone g-FW⁻¹); however, overall total carvone ((mg carvone g-FW⁻¹) × g culture FW) increased because of the production of greater biomass obtained per vessel.     

Opposite effects of exogenous sucrose on growth, photosynthesis and carbon metabolism of in vitro plantlets of tomato (L. esculentum Mill.) grown under two levels of irradiances and CO2 concentration

The long-term effects of exogenous sucrose (3 percnt;) on growth, photosynthesis and carbon metabolism ofin vitro tomato plantlets were investigated under two sets of growth conditions that respectively favor source- or sink-limitations of photosynthesis: 1) low photosynthetic photon flux (PPF) (50 μmol m⁻² · s⁻¹) and low CO₂ concentration (400 μmol mol⁻¹) and 2) high PPF (500 μmol m⁻² · s⁻¹ and high CO₂ concentration (4000 μmol mol⁻¹). The supply of sucrose under source-limitation conditions increased the growth, the maximal photosynthetic rate, the chl content, the maximal quantum yield of Photosystem II estimated by the Fv/Fm chl fluorescence ratio as well as the soluble sugars (hexoses, sucrose) and starch contents in roots, young and mature leaves when compared to those of photo-autotrophic plantlets. Also, sucrose feeding under these conditions strongly increased the activity of sucrose synthase (SS) (EC 2.4.1.13) in roots and young leaves whereas the activities of sucrose phosphate synthase (SPS) (EC 2.4.1.14), acid invertase (INV) (EC 3.2.1.26) and ADP-glucose pyrophosphorylase (ADPGppase) (EC 2.7.7.27) were highly stimulated in roots and mature leaves. Contrary to these observations, the supply of sucrose to plantlets developed under high PPF and CO₂ concentration decreased growth and led to a somewhat lower maximal photosynthetic rate relative to photo-autotrophic plantlets. These negative responses to exogenous sucrose were accompanied by stronger accumulations of hexose and starch, larger stimulation of INV in mature leaves developed under conditions of sink limitation than those from source limitation conditions. Moreover, under high PPF and high CO₂ concentration, exogenous sucrose led to a marked repression of the SPS activity and caused much lower stimulations of ADPGppase in mature leaves than those observed at low PPF and low CO₂ concentration. We therefore conclude that under our experimental conditions, the interactive effects of exogenous sucrose and environmental conditions on growth and photosynthesis could be rationalized by the source-sink equilibrium of thein vitro tomato plantlets.     

Blue and red light-emitting diodes with or without sucrose and ventilation affect in vitro Growth ofRehmannia glutinosa plantlets

Single-node, in vitro cuttings ofRehmannia glutinosa were transplanted to MS basal media and grown for 30 d. Plantlets were grown under various culture conditions: four different light qualities (red LEDs, blue LEDs, mixed LEDs, and fluorescent); with sucrose (30 mg.L^-1) or without (0 mg.L^-1); with air exchanges (3.5 h.^-1) or without (0.1 h.L^-1). Highest dry weights were obtained from plantlets under blue LEDs with 3.5h.L^-1 air exchanges. Light source did not affect shoot elongation in ventilated conditions, but without ventilation, the shoots of plantlets under red LEDs were twice as long as for plantlets growing under other types of lighting. Plantlets grown without sucrose showed little difference in photosynthesis under any of the tested light qualities. In contrast, the photosynthetic rate of those in the sucrose-containing media varied according to light source.     

Physiology of effects of temporary immersion bioreactors on micropropagated pineapple plantlets

Summary Temporary immersion bioreactors are an efficient tool for plant mass propagation because they increase multiplication rate and plant quality. Little knowledge is available on the ecosystem and physiological behavior of plantlets when using this new culture technique. In order to evaluate the effects of the conditions on physiological change of pineapple plantlets, a factorial experiment was conducted, where axillary clusters were cultured under two levels of photosynthetic photon flux (PPF): 30 μmol m⁻²s⁻¹ (low) and 225 μmol m⁻²s⁻¹ (high), using two culture methods (conventional micropropagation in liquid medium and a temporary immersion bioreactor) during the elongation phase. CO₂ concentration in the headspace volume container was measured during a whole cycle of temporary immersion (3h). At the time before the next immersion period, the levels of CO₂ increased significantly to 14171 μmol mol⁻¹ at high PPF. The maximal photosynthetic rate as well as the maximum quantum yield of photosystem II were low for plantlets cultivated in the femporary immersion bioreactor at high PPF. However, these plantlets showed large increases in sugar and nitrogen uptake and also increases in dry weight and foliar area. These results indicate that shoot growth did not totally depend on the photosynthesis process. In vitro pineapple plantlets appeared to use more nutrients in the culture medium than those from photosynthesis. In summary, temporary immersion bioreactor-derived plantlets showed remarkable nutrient uptake, indicating a higher photo-mixotrophic metabolism.     

<Emphasis Type="Italic">In vitro</Emphasis> fruiting and seed set of <Emphasis Type="Italic">Capsicum annuum</Emphasis> L. CV. Sweet banana

Summary Plantlets of Capsicum annuum L. ev. Sweet Banana regenerated via somatic embryogenesis from immature zygotic embryos were capable of producing flower, fruit, and seed when cultured in small tissue culture containers. In vitro floral buds were first formed on plantlets that grew on plantlet development medium [agar-gelled Murashige and Skoog (MS) basal medium containing 1 mgl⁻¹ (5.3 μM) α-naphthaleneacetic acid (NAA)] in a growth room at 22°C and continuous illumination. However, floral buds rarely developed further into mature flowers. This problem was overcome using the vented autoclavable plant tissue culture containers. In vitro fruit formation and ripening was observed when liquid half-strength MS basal medium supplemented with 5 μg ml⁻¹ silver thiosulfate, 1 mg l⁻¹ (5.3 μM) NAA, and 3% sucrose was added to the surface of the plantlet development medium. Hand-pollination improved fruit set. Further research in needed to determine why the pepper seeds formed in vitro failed to germinate.     

Photosynthetic characteristics of Cymbidium plantlet in vitro

The photosynthetic characteristics of the Cymbidium plantlet in vitro cultured on Hyponex-agar medium with 2% sucrose were determined based on the measurements of CO₂ concentration inside and outside of the culture vessels. The CO₂ measurements were made with a gas chromatograph at a PPF (photosynthetic photon flux) of 35, 102 and 226 mol m^-2 s^-1, a chamber air temperature of 15, 25 and 35°C and a CO₂ concentration outside the vessel of approximately 350, 1100 and 3000 ppm. The net photosynthetic rates were determined on individual plantlets and were expressed on a dry weight basis. The steady-state CO₂ concentration during the photoperiod was lower inside the vessel than outside the vessel at any PPF greater than 35 mol m^-2s^-1 and at any chamber air temperature. The photosynthetic response curves relating the net photosynthetic rate, PPF, and CO₂ concentration in the vessel and chamber air temperature were similar to those for Cymbidium plants grown outside and other C₃ plants grown outside under shade. The results indicate that CO₂ enrichment for the plantlets in vitro at a relatively high PPF would promote photosynthesis and hence the growth of chlorophyllous shoots/plantlets in vitro and that the plantlets in vitro would make photoautotrophic growth under environmental conditions favorable for photosynthesis.Abbreviations C_in CO₂ concentration in the culture vessel - C_out CO₂ concentration outside the vessel (in the culture room) - PPF photosynthetic photon flux     

Plantlets from encapsulated micropropagated buds of M.26 apple rootstock

In order to be considered usable as synthetic seeds, encapsulated explants sown underin vitro orex vitro conditions must be able to produce whole plantlets. Ninety percent of non-encapsulated M.26 apple rootstock single nodes produced a plantlet (i.e., a well-formed shoot with a root system) after 30 days of culturein vitro if the explants were previously given a 24-hour treatment with 24.6 μM IBA and 15 g 1⁻¹ sucrose in darkness. In contrast, when the single nodes were encapsulated in a calcium-sodium alginate bead immediately after the same treatment only 10% of the encapsulated explants formed a plantlet. Addition of growth regulators to the artificial endosperm and culture of the single nodes for root primordia initiation for 3, 6 or 9 days in darkness before encapsulation allowed production of 58%, 60% and 66% of plantlets, respectively.     

Effect of sucrose application,minerals, and irradiance on the <Emphasis Type="Italic">in vitro</Emphasis> growth of <Emphasis Type="Italic">Cistus incanus</Emphasis> seedlings and plantlets

To study the effect of sucrose on the sink-source relationship in in vitro-grown plants, Cistus incanus seedlings and plantlets were grown horizontally in a two-compartment Petri dish (split dish), with the root system in one compartment and the shoot in the other. Shoots and roots were exposed to different sucrose concentrations (0–30 g dm⁻³), two irradiance levels (25 and 160 μmol m⁻²s⁻¹) and the presence or absence of a minimum medium containing minerals and vitamins (M medium). Root and shoot biomass of the seedlings was enhanced by an increase in irradiance when the growth medium was not supplemented with sucrose indicating the role of photosynthesis in biomass production. When sucrose was added to either organ growth was enhanced as well. In the presence of sucrose in the root compartment, sucrose applied to the shoot compartment enhanced growth of both organs under low irradiance, while under high irradiance, sucrose had no further additive effect. In the absence of sucrose in the root compartment, the enhancement of root biomass by sucrose added to the shoot compartment was lower under high irradiance than under low irradiance. The response of Cistus plantlets to sucrose and irradiance differed from that of seedlings, probably reflecting a greater susceptibility of the plantlets to sucrose feedback inhibition on photosynthesis and biomass accumulation. The decrease in root and shoot growth when M medium was added to the shoot compartment and the relatively better growth of these organs when the roots were supplied with minerals and the shoot with sucrose, indicate that growth of the two organs in our experimental set-up was regulated by opposing fluxes of C and nutrients.     

Influence of nitrogen and phosphorus on induction embryogenic callus of sorghum

Crown and leaf slices of in vitro plantlets of a non-flowering Vetiveria zizanioides from Java were used to induce compact calli and to regenerate plantlets. The influence of different growth regulators (2,4-dichlorophenoxy acetic acid, 6-benzylaminopurine), sucrose concentrations (10–100 g l⁻¹), cultivation in light or dark, and cultivation time on callus induction medium (6 or 12 weeks), on the induction of compact callus and the subsequent regeneration of plantlets was studied. Up to 75% of crown slices cultured on modified Murashige and Skoog medium supplemented with 2.26 μM 2,4-dichlorophenoxy acetic acid, 2.22 μM 6-benzylaminopurine and 75 g l⁻¹ sucrose developed compact callus. For subsequent regeneration of plantlets, callus induction in the light for 6 weeks on the callus induction medium containing 10 g l⁻¹sucrose, and subsequent transfer to the regeneration medium, was the best procedure, regenerating plantlets on around 60% of the crown or leaf slices, with up to 100 plantlets per slice. We have compared the efficiency of the above mentioned procedure with several other methods to regenerate plantlets. Our findings indicate that the procedure developed in this study was best in regenerating plantlets for the used vetiver variant. This revised version was published online in June 2006 with corrections to the Cover Date.     

Similarity of growth patterns between plantlets and seedlings of Brassica campestris L. under different in vitro environmental conditions

Explants and seeds of Brassica campestris L. were cultured on Murashige & Skoog (1962) medium with and without sucrose in a vessel with different numbers of air changes per hour under different PPF (photosynthetic photon flux) conditions. The growth and development of plantlets in the vessel were similar to those of seedlings when cultured under the same in vitro environmental conditions. The growth and development of seedlings when cultured under the same in vitro environmental conditions. The growth and development of plantlets/seedlings were greater for treatments with a higher number of air changes per hour and a higher PPF regardless of the sucrose concentration in the culture medium.The CO₂ concentration in the vessel with a lower number of air changes per hour decreased to approximately 100 ppm during the photoperiod on day 21 due to the photosynthetic activities of the plantlets/seedlings. The low CO₂ concentration, in turn, reduced the net photosynthetic rate of plantlets/seedlings in the vessel, and thus affected their growth and development.Abbreviations C_in CO₂ concentration in the culture vessel - C_out CO₂ concentration in the culture room - MS mineral composition of Murashige & Skoog (1962) medium - PPF photosynthetic photon flux     

Avocado shoot culture, plantlet development and net CO2 assimilation in an ambient and CO2 enhanced environment

Summary The proliferation and survival of avocado nodal cultures of juvenile origin were affected by the form and concentration of nitrogen. Optimum growth was achieved on modified Murashige and Skoog medium containing 67% KNO₃ and 33% NH₄NO₃ with total N of 40 mM supplemented with 100 mg l⁻¹ myo-inositol, 1 mg l⁻¹ thiamine HCl, 30 g l⁻¹ sucrose, and 4.44 μM BA with a 16-h photoperiod (120–150 μmol m⁻² s⁻¹). Proliferating shoots and plantlets were photosynthetically active. Better shoot growth and accumulation of higher biomass occurred in a CO₂-enriched environment than under ambient CO₂ conditions. CO₂ assimilation efficiency, however, was higher under the latter conditions than in a CO₂-enhanced environment, e.g., 31±7 and 17±2 μmol CO₂ m⁻² s⁻¹, respectively. The net CO₂ assimilation rates of in vitro grown plantlets were comparable to those of seedlings ex vitro.